1 Introduction

2 Features

2.1 Subsetting
2.2 Color Coding
2.3 Surfaces
2.4 Electrostatics
2.5 Distance Calculations
2.6 Maps
2.7 Atoms
2.8 Bonds
2.9 Contours
2.10 Colors
2.11 Simple Property Mathematics
2.12 Objects
2.13 Pair-Wise Interaction (Matrix) Representations
2.14 Stereo/Split Screen Operations

3 Getting Started

3.1 Grasp Environment
3.2 Running Grasp
3.3 Dial and Mouse Movement
3.4 Data Files
3.5 The Textport
3.6 Exit

4 Menus

4.1 Display Menu
4.2 Build Menu
4.3 Calculate Menu
4.4 Mouse Functions Menu
4.5 Read Menu
4.6 Write Menu
4.7 Formal Subset Menu
4.8 Programs Menu
4.9 Set Parameters Menu
4.10 Miscellaneous Menu
4.11 Help Menu
4.12 Macros Menu
4.13 Quit GRASP

5 Command Line

5.1 Subsetting Codes
5.1.1 Negation, Ranges, and Concatenation in Subsetting Codes
5.1.2 Atom Subsetting Codes
5.1.3 Surface Subsetting Codes
5.1.4 Bond Subsetting Codes
5.1.5 Backbone Box Subsetting Codes
5.1.6 Pair-Wise Interaction (Matrix) Strand Subsetting Codes
5.2 Coloring
5.3 Precise Rotation and Translation
5.4 Listing Atom Properties
5.5 Altering Radii and Charges
5.6 History and ! Commands

6 Control Keys

7 Worked Examples

7.1 Color a molecular surface by electrostatic potential
7.2 Display surface potential and curvature side by side
7.3 Surface two interacting parts of a molecule and select the interface
7.4 Calculate the occluded accessible surface area between these parts
7.5 Display atomic B­values on a surface
7.6 Calculate and display effective dielectric from a single charge site
7.7 Calculate surface area by hydro-phobic and -philic residue
7.8 Find and display all residues within 3 Angstroms of an active site
7.9 Find the common volume between two superimposed molecules
7.10 Form a 6-helix bundle from a single helix

8 Future Developments

8.1 General Improvements
8.2 Intelligent DelPhi
8.3 Secondary Structure Display
8.4 Docking with Realistic Energies

Appendix A File Formats

A.1 Grasp Surface File
A.2 DelPhi Potential Map
A.3 DelPhi Charge File
A.4 DelPhi Radius File
A.5 Protein Data Bank (PDB) File and Grasp Variants
A.6 Grasp Property File
A.7 Grasp Script File
A.8 Grasp Macro File
A.9 Pair-Wise Interaction (Matrix) Energy File

Appendix B .init_grasp Commands

Appendix C Grasp Data Files

Addendum for Version 1.2


1 - Introduction (By Anthony Nicholls)

Grasp came about because I wanted to visualize electrostatic potentials at surfaces, in particular the surface of biological molecules. Barry Honig's lab is well known for the program DelPhi which calculates electrostatic potentials from the Poisson­Boltzmann equation, and has the led the way in applying this equation in structural biology. The program can be used to get productive quantitative numbers for a variety of biochemical phenomena but qualitative visualization had been limited to isopotential contouring, typically with a program such as Insight from Biosym Technologies. The limitation of such an approach is that such contours do not capture local topology or shape. They often extend significant distances away from the molecule while one would expect most of the 'action' to be close to the molecule, in fact at the surface of molecules. So I decided it was time to attempt a graphics program which emphasizes surfaces and electrostatics.

I would not have been so confident in starting this project had it not been for considerable groundwork laid by others, in particular Kim Sharp. He and Mike Gilson had devised a novel algorithm some years ago for calculating the molecular volume (i.e. water inaccessible volume) using a cubic lattice. This method, though not efficient when the lattice spacing is small relative to atomic radii, can be made rapid at lower resolution. Given the molecular volume, Kim had also reinvented a technique commonly known as the "Marching Cubes" algorithm to produce a surface tessellation. Putting these together in an optimized form produced a rough and ready surface description which could be easily visualized on a Silicon Graphics Iris (SGI) computer. The potentials at surface points could then be interpolated from the 3­D map produced by Delphi and color coded to indicate the result.

The initial results were surprisingly good, both in terms of aesthetics and usefulness. The large difference in dielectric between water and the interior of proteins modeled by DelPhi means that local electrostatic effects can dominate global ones. So, for instance, an active site can be negative even when the protein total net charge is very positive. This is seldom seen if Coulomb's Law is used to calculate potentials because of the long range nature of (1/r). Grasp was immediately able to display this consequence of electrostatic screening, for instance showing the deeply negative binding site of the catalytic magnesium of RNAse H, crystallized without that ion by Yang and Hendrickson.

So it was clear that this approach held some merit. The combination of surface shape and electrostatic potential was synergistic. Moreover I began to see that just having a rapid surfacing and visualization algorithm was useful. For instance it was simple using surface connectivity to display only the internal cavities of proteins. Here the initial use was the bacteriorhodopsin structure of Henderson which has numerous "holes" surrounding the retinal moiety.

It was also instructive to "project" properties of the underlying atoms onto the surface and color code them, an example being the B values normally accompanying crystallographic structures. So I began to see the surface itself as useful construct, regardless of electrostatics. This became a central tenet of Grasp, that surfaces and atoms should be treated with equal importance.

There was still a need for other electrostatic representations, such as isopotential contours, projection planes, field lines, etc. Also, since typical use of the program was visualization of large molecules with thousands of atoms, I devised a simple representation for those atoms that was fast to display and could be colored by property. This led to other representations of atoms and groups of atoms.

The program grew beyond my initial plans. Hopefully, however, the program maintains a coherent philosophy. For instance the use of surfaces both for displaying properties and as objects in their own right, the visualization of electrostatic properties, and more lately the generalization of the idea of an object representing both a set of atoms (as the surface does) and a property (such as electrostatic potential). An example of the latter is the DNA representational project of Rex Bharadwaj. Here DNA bases can be represented as elongated boxes whose width can represent a property associated with that base, such as base twisting, sliding or rolling.

The program has achieved most of the goals I had concerning the development of these ideas. Of course in their actualization they have spawned many more. But hopefully the present version is at least complete enough to be useful.

Comments as always are much appreciated.

Anthony Nicholls

October 1992

2 - Features

Grasp uses a perspective-based view, which means things farther away from the eye are smaller. To enlarge a view, one simply moves it closer to the eye. Manipulations are by mouse or by dials. Molecules are mapped to a "unit box" which can be displayed with the front side removed. There are also embedded cross­hairs to remind the user of any rotations and translations they have applied. The default clipping planes are very close to the "eye" position and very far away. These can be altered via a slice control tool (section xxx). The background is either black or that produced by the unit box. The default window size may be changed in the usual window resizing manner. There is also a fullscreen option where the entire screen is used by the display. All functions are accessed by hierarchical menus via the right mouse button, or via the command line (section 5). Commands may also be read in from a script file (section A.7). All display objects (surfaces, atoms etc.) are independent - they can all appear at the same time or be individually hidden from view.

2.1 - Subsetting

A central feature of Grasp is the ability to specify a subset of atoms, surface vertices, bonds, objects, etc., based upon a very wide range of properties.

The first method of selecting a subset is by typing property subsetting codes on the command line or in response to a menu query. Properties which can be used for atoms are atom name, atom number, residue name, residue number, residue projection, chain name, charge, radius, potential, original coordinates, screen coordinates, general properties 1 and 2 (which can be assigned any number desired), molecule number, accessible area, distance, formal subset name, and discrete atom color. Properties for surfaces are potential, original coordinates, screen coordinates, general properties 1 and 2, distance, formal subset name, surface number, curvature, vertex number, and discrete vertex color. There is a deliberate similarity between these two lists based on the program philosophy of equality between surfaces and atoms.

The general property fields can be used in a variety of ways. For instance any atom property can be mapped to a surface and stored (and hence displayed) as general property 1 or 2. Or the user can produce a new quantity out of others by the simple property mathematics utility. Or properties can be imported.

Variables are either characters or numbers. The latter may be specified as ranges, for instance one can select all atoms which have a residue number between 5 and 10 or a charge greater than zero. Character variables (except formal subset names) may include wild cards, so one can select all carbon atoms or all carbon atoms which have the character "1" in the third position of the atom name. There are also short cut names for atoms in a protein backbone or in side chains, and for residues which are ionizable, hydrophobic or hydrophilic. All specifications can also be negated, so that one can select all atoms that are NOT carbon atoms.

The purpose of subsetting depends upon the context. One of the simplest uses is to alter the display color of a subset. For instance, one might want to employ a color scheme which displays all positively charged atoms one color and all negatively charged atoms another, or all hydrophobic residues yellow and all hydrophilic purple, etc. Colors are specified by the index number (0­99) assigned to each color in Grasp.

Another use of subsetting is creating a formal subset. Formal subsets are assigned names, either by the program, in which case a hierarchical naming convention is used, or by the user. Formal subsets may be spatially manipulated independently of the rest of the surfaces/atoms. They can also be referred to by name in all subsequent operations. Formal subsets may be sets of atoms or portions or collections of surfaces, or may have mixed character - a surface may be associated with a set of atoms, or a set of atoms associated with a surface. For instance, as well as selecting an active site surface one might want to associate all atoms which are in contact with that surface.

Most other subsetting uses depend on the action being undertaken. For instance, when the surfacing subprogram is activated the user has the option to enter a subset of atoms. One could surface a single helix in a protein.

Most subsetting codes are exclusively AND based - subsetting by progressive refinement based upon properties. The reason for this is that one of the most useful applications of Grasp is using it to look for correlation. So one might want find all residues which are a certain distance from the molecular surface AND charged.

The second method of selecting a subset is "scribing" the surface. This is under menu control and allows the user to "draw" upon a surface the border of a region of interest with the mouse cursor in much the same way in which one might with a Macintosh­like draw program.

2.2 - Color Coding

Grasp supports two different modes of color coding.

The first color coding mode, 3 color continuous, requires three numerical values, called "control" values, along with three colors, one color per value (colors are defined by RGB triplets, or by an index into a list of colors). Color coding is then implemented as follows: If a number is less than the minimum of the three control values it is assigned the color associated with that minimum value. If it is greater than the maximum value it is assigned that value's color. If it is between the minimum and middle values the assigned color is found by linearly interpolating between the minimum and middle colors, and if it is between the middle and maximum values by linearly interpolating between the middle and maximum colors. By linearly interpolating is meant the following: Colors are made up of red, green and blue components, each component having a strength of 0.0 to 1.0. If a number is, for example, halfway between one value and another, then its interpolated color is similarly halfway between the two colors assigned to those values, i.e. its red, blue and green components are half those of one color and half of the other.

Grasp also supports 2 color continuous mode which is equivalent to 3 color continuous mode but with the middle value and color set to be the same as that of the minimum value and color.

Continuous color maximum, minimum and middle values may also be adjusted via a mouse activated widget. This is useful in rescaling the color code to bring out particular features on the fly. Independent widgets appear for each continuous property displayed. These also offer access to other features via drop-down menus.

The second color coding mode is zonal coloring, wherein a certain range of values is assigned a certain color. The color boundaries between different zones are sharp. This is also referred to as discrete coloring. Default colors are provided for all properties - for electrostatic potentials they are red, white and blue, with red for the minimum value (typically negative), white at zero, and blue for the maximum value (typically positive). However these colors may be also set by the user. Default maximum and minimum control values are taken as the maximum and minimum values of the property being represented (surface potential, atom distances, etc). The middle value is set to zero unless the maximum and minimum are both greater than or less than zero, in which case it is set to the average of those two values. These control values can also be explicitly altered.

2.3 - Surfaces

Grasp supports two types of surfaces, molecular and accessible. The molecular surface is defined as the boundary of that volume within any probe sphere (meant to represent a water molecule) of given radius sharing no volume with the hard sphere atoms which make up the molecule. The accessible surface can be defined as the locus of the centers of all possible such probes described above in contact with the hard sphere atoms. Alternatively it can be defined as the hard sphere surface if each atomic radius is increased by the probe radius. The default probe radius for each type of surface is 1.4 Angstroms. This can be changed by the user.

Everything which can be done with molecular surfaces can be done with accessible surfaces. Therefore except where differentiated they will both be referred to as molecular surfaces. The surfacing resolution, the lattice spacing used to generate the surface, is determined automatically. The lattice spacing used in the process is scaled relative to the largest dimension of the molecule, hence coarser lattices are used for larger molecules. This scaling has the advantage that the surface of very large molecules can be as easy to manipulate as that of small molecules. Though the surface of larger molecules will then be less accurate, one is often interested in coarser features of such molecules.

If the molecule has only a few atoms, this method can lead to a lattice spacing at which the method Grasp uses is inefficient, hence there is a minimum allowed lattice spacing. This parameter can be set by the user to force use of a coarser grid (as might be preferred to improve draw speed or to overcome memory limitations).

Surfaces can be constructed for all atoms or for subsets of atoms. The process takes a few seconds at most and results in a smooth tessellation of the surface which is colored white but shaded by the SGI lighting routines. This can be quite slow on some machines as there may typically be 20,000 triangles. To enhance drawing speed there are three other draw modes. The first is a rendered surface (the triangles are filled in) but the lighting calculations are simplified (pseudo­lit) and done in software rather than by using the SGI hardware calls. The second is a mesh representation, where the triangles are not filled in. The third is a points­only representation. The default surface produced by the program can be preset in the .init_Grasp file (see appendix B).

All displays of surfaces (and also contours, atoms and bonds) are depth shaded. This means that the farther away a part of the surface the darker the color. Its color is interpolated to black based on the distance from the viewer. Grasp uses a dynamic depth range, where the front edge, where interpolation begins, is determined by the nearest point on the structure to the viewer, and the back edge, the depth at which the color is set to black, is a fixed distance behind this point. Depth cueing makes a dramatic difference to the mesh representation, and to a lesser extent to other representations. Depth cueing parameters can be set by the user.

For the lit and pseudo­lit surfaces the direction of the incident light may be changed. Other material properties of the surface such as reflectivity, transparency, etc., are not currently user­configurable.

Surfaces are colored by assigning colors to each vertex. This can be done according to any value associated with that vertex. Alternatively, surfaces can be colored by selecting a subset and assigning a discrete color. There are many ways of selecting a surface subset based on vertex properties, associated atom properties, or by scribing. Any surface or subset can be "uncolored" i.e. undisplayed. Hence one can remove portions of surfaces to create "windows" into the underlying molecule. Any surface property can be interrogated using the mouse buttons - placing the mouse at any given point and clicking returns a value or values associated with the nearest surface vertex. Which property values are returned, and by which mouse button, can be set by the user.

Surface properties can be classified as those which can be used for subsetting and displayed, and those which can be used for subsetting but not displayed. Surface properties which can be displayed are generally calculated within the program and include electrostatic potential, curvature, distance (to another surface or set of atoms), and the two general property fields. The general property fields can be assigned other property values to make possible the display of properties which would otherwise be undisplayable. Surface properties which cannot be displayed are original coordinates, screen coordinates, discrete vertex color, surface number, formal subset name, and vertex number.

Surface potentials are interpolated from potential maps. Distances as a surface property are calculated either from another surface or surface subset, or from a set of atoms, and in each case are the minimum such distances. Curvature is as defined in Nicholls et al and is derived from a concept of local hydrophobicity. Briefly, each possible placement of a water "sphere" against the surface of the molecule reduces the accessibility of that water to other waters. Against a concave surface this accessibility is less than against a convex surface, and a formal correspondence can be made between contact to an arbitrarily complicated surface to contact with a sphere of a certain curvature. Curvature thus defined is a property of the accessible surface, but can be uniquely mapped to the molecular surface. When color coded it reinforces the effect of SGI lighting in that surface hollows are made distinct from surface projections, and so is useful in visualizing patterns of surface shape.

Surface data (which means vertex coordinates, connections, and normals), and any properties, calculated or assigned, of any surface or subset of surfaces can be written to a data file. These data files can be read in during program execution or at startup. Surface data can be appended to other files to make surface "libraries". Since subsets of surfaces can be saved one could, for instance, make a library of the surfaces of the active sites of different enzymes.

The property data of a surface or subset can also be saved as an ascii file for analysis, or for temporary storage. This file can also be read back in as a user-generated function mapped to a particular surface.

Surfaces or subsets of surfaces can have their surface normals inverted. In this way one can look at molecules from the inside. One can also do like­with­like comparisons of two surfaces which might form complexes by inverting one surface of the pair. Surface properties can also be inverted, for instance positive turned to negative and vice versa through the simple property mathematics utility.

Along with cavity surfaces (which can be thought of as a surface subset) and contour surfaces, one can calculate the area of any molecular surface or subset of a molecular surface, and similarly the volume enclosed (noting that the volume is not meaningful if the surface is not closed). The surface area for an accessible surface gives a measure which has often been associated with hydrophobicity, since it is related to the number of water molecules in contact with the molecule. Grasp also has a more accurate surface area subroutine for atom by atom accessible area.

Any surface or subset can also be attached to the rotation and translation dials alone. This creation of a formal subset can then be manipulated independently. This allows for parts of surfaces to be removed, compared, docked, etc.

2.4 - Electrostatics

Grasp includes a Poisson­Boltzmann (PB) solver which is a similar but simpler version of that used by DelPhi. The fields calculated by it are for qualitative use only. For quantitative use there is full support for the output from DelPhi, in terms of potential maps, dielectric maps, modified pdb files, charge files, size files, etc. Grasp does not yet contain an interface to DelPhi, hence that program has to be run separately.

The Grasp PB solver uses two 33 cubed grids, one nested within the other. The inner grid dimension is set to be larger, by the diameter of one water molecule, than the maximum x, y or z dimension of the collection of atoms used in the calculation. The second grid is twice the size of the first, with the same center. The potentials on the outer grid are solved for first, then interpolated and refined further on the inner grid. Potentials are then interpolated to a 65 cubed grid the same size as the outer grid. This final grid, or "map" as it is referred to, is then used in all subsequent calculations. It may also be written out in DelPhi form.

Although there is no choice in the sizing of these grids, the user has control of the inner and outer dielectric constants, the probe radius used to determine water inaccessibility, the salt concentration, and the ion exclusion radius. There is no support for the nonlinear equation, for periodic boundary conditions, membrane slabs and holes, or any other DelPhi features.

Once a map is calculated, it can be evaluated in several ways. Isopotential (also referred to as "through space") contours may be calculated at any value, given any color, and displayed as solid surfaces, meshes or points. Potentials may be interpolated at any molecular surface, and at any set of atoms (trilinear interpolation is used throughout). The electric fields may be calculated at a set of points and represented in magnitude and direction as three dimensional arrows. Molecular dipoles may be calculated and similarly displayed. Field lines can be calculated from a set of points, colored and displayed in 1­D or 3­D (lines or tubes). The potential may also be interpolated at a slice plane perpendicular to the Z direction (parallel with the screen). This latter display is updated as the map/molecule is moved, or alternatively as the position of the slice plane is altered.

Values at surfaces and atoms may be colored by 2 or 3 color continuous or by discrete colors. The Z plane will only show the former. Field vectors may have their magnitude encoded in their length. Field lines can be assigned directionality and color when calculated.

2.5 - Distance Calculations

Grasp will calculate minimal distances from surfaces to surfaces, from surfaces to atoms, from atoms to surfaces and from atoms to atoms. In the case of atoms there is also the option to subtract the assigned van der Waals radii from the distance.

An example of a novel use of distances in Grasp is to calculate a "depth" map, i.e. the depth of atoms from the surface to every atom. Distance maps are also useful in defining interfaces between domains, either surface­wise or atom­wise.

2.6 - Maps

Grasp contains room for two internal "maps", i.e. 65 cubed, cubic lattices. Internally generated maps are stored in the first of these arrays. Maps read in are put in the second array. Simple operations are allowed on and between maps, such as differences, sums, etc. Maps can also be swapped so that both can be internally generated or both externally generated by DelPhi. Difference maps are particularly useful to highlight the effects of changing parameters such as charge, radii, salt concentration, dielectrics, etc.

As one of the three primary external data files supported by Grasp (the other two being protein data bank (.pdb) files for atoms and surface representation files (.srf) for surfaces), maps may be read at startup, so that one can analyse maps without any atom data or surface data. Although maps are usually associated with electric potential, they can be used quite generally for any 3­D data, though at present Grasp requires the grid to be 65 cubed. For example, the consensus volume option in Grasp, finding the common volume between a set of molecules, results in values assigned to a 65 cubed grid which can then be manipulated and displayed as a potential map (e.g. Z­plane projection, isopotential contours). The dielectric boundary map or salt exclusion map from DelPhi can also be read in as this form.

2.7 - Atoms

Atomic coordinates are the fundamental data structure from which everything else is derived. Grasp does not contain any "build" utility and hence is dependent on external files for this data. Primary support is for PDB files, the standard crystallographic format, along with certain variants.

Atoms can be displayed in several ways. The traditional method, which is included in Grasp, is as spheres of a given radius. This is often referred to as CPK modelling. In Grasp the surfaces of the spheres are lit. CPK can be demanding on the graphics resources for large molecules. An efficient alternative is to represent the atom as a circle of the correct radius always oriented flat with respect to the viewer. With a little differential coloring these flat circles can be given an apparent three­dimensionality. There is also an option to color these circles with patterns. There is another representation which gives small, uniformly sized circles for use with bond representations. Spheres may also be drawn with lines or dots.

Atoms can be colored discretely - any subset of atoms can be assigned any color, or continuous color colding may be used. The properties supported for this are potential, distance, charge, and general properties 1 and 2. Atoms can be uncolored to remove them from view. Atom colors are depth shaded.

Upon reading a PDB file, radii are set to default values from an external file (which the user may edit). This file is in the same format as the control file for atom size used by DelPhi. Similar files may be read during program execution. Charges are 0 by default when a structure is read, but Grasp can read DelPhi charge files and assign charges based on the descriptions therein. Some sample files are provided, such as those to assign charges to each ionizable residue of a protein. Radii and charges may also be assigned via the command line by specifying a radius/charge and the subset of atoms to have this radius/charge. Keyboard commands also support intrinsic operations such as multiplication of radii/charges by constants, or addition of constants.

Since charges are only of importance for electrostatic purposes and since charge is also a display property, it may be utilized as a "dummy" variable, and actually represent another physical property. This becomes particularly useful when combined with the DelPhi control file format. For instance if one is interested in a per­residue property, say helix­forming propensity, a control file can be constructed with one line for each residue and its property value. This can then be read into Grasp, the atoms of each residue will be assigned a "charge" equal to that residue's helix­forming propensity, which can then be color coded and displayed.

Grasp also supports three variants on the standard PDB file which involve the fields to the right of the atom coordinates. These typically contain occupancy and B­values. One option is to read this information in as general properties 1 and 2. There is also an option to read these fields in as the radius and charge of each atom because this is what they are used for in DelPhi modified PDB files. Finally, for higher precision, the entire field to the right of the coordinates can be read, in free format, as general properties 1 and 2. Files in the these formats can all be written from within Grasp.

Separate molecules will be recognized from within a single PDB file if separated by "TER" statements. Each will be assigned an index, i.e. a molecule number, which as a property is analogous to "surface number" in Grasp. Molecules can be superimposed using the Kabsch algorithm, which gives the best rotation and translation (RMS­wise) between molecules or parts of molecules. The only restriction is that the same number of atoms from each molecule must be used to determine the minimum RMS difference. Grasp will not yet superimpose surfaces.

Grasp contains algorithms for calculating both the volume and surface area of molecules or subsets of atoms. Surface area can be either accessible area or that of the van der Waals surface. Control is given to the user over the precision of these calculations.

Grasp does not contain methods to alter structures such as torsional rotations, minimization, etc., with one key exception. Grasp allows independent rotations and translations of defined subsets, which may then be "fixed" relative to each other. For instance, a substrate may be selected and moved relative to an active site. Upon making the transformations, new surfaces, distances, electrostatic fields, etc., can be calculated based on the new coordinates. One can undo transformations which have not yet been "fixed".

Atom properties can be queried in the same way as surface properties, i.e. point and click. Atom properties can even be queried when covered by surfaces. The "atom picking" function can also be set to report geometric parameters such as distance, angle, and torsion angle between picked pairs, triplets, and quadruplets of atoms.

2.8 - Bonds

Bonding patterns are calculated upon reading in a pdb file. Bonds may be represented in three ways: lines, sticks, and cylinders. Lines are the traditional line drawings used by most programs. Cylinders are a three dimensional variant of this, the diameter of which can be set by the user. Sticks follow the method of Kuznetsov and Lim in which bonds are represented by quadrilateral tubes. The advantages of this approach are that the bonds are made significantly more three dimensional, inter bond angles are well brought out, and the display is relatively quick to draw.

Bonds can be colored based upon a preset pattern, upon transferring the discrete colors of the underlying atoms, or by subsetting based on the properties of those atoms. They can also be selectively undisplayed by being "uncolored". Bond colors are depth shaded. Properties of atoms can be queried by picking at bond ends.

2.9 - Contours

Isopotential contour surfaces can be constructed at any potential value for either internal map. Contours can be displayed in all the surface modes available to molecular surfaces: lit, pseudo­lit, mesh and points. They are also independently depth-shaded. Contours are not automatically recalculated if the parent map changes, though contours can be deleted and recalculated. Volume and surface areas may be calculated for any contour.

2.10 - Colors

Grasp supports 99 independent indexed colors. These can be set during program execution using a color palette, or in an external file as RGB triplets. All changes to colors are automatically saved (except for colors 91-99, which are always equal to the default colors 1­9), thus the user can design their own set of colors or use those provided. Color 0 is always black and is also used as a flag to prevent display of that an object - an atom colored 0 is hidden. This "uncoloring" applies to surfaces, bonds, backbone boxes and matrix strands. Grasp also has undo and restore commands for atoms and vertices. Undo removes the previous coloring, while restore acts on objects colored 0 by giving them the color previously assigned. Once a color is assigned to an object it becomes a property which can be used in subsequent subsetting selections.

The use of assigned color as a property allows for some quite flexible subsetting. For example it can act as a bridge between atomic and surface properties. Suppose we want to find all vertices of a surface which are concave (have a calculated curvature less than zero)and which are formed by hydrophobic residues. One way to do this is to color all hydrophobic residues one color, transfer that color to the surface, then select all vertices which have that color AND which have curvature less than zero.

Color can be used to build up subsets based upon disparate properties which could not be specified in a single command. For instance we can select all atoms which are in a region of positive potential and belong to negatively charged residues plus all those atoms which are negative but belong to positively charged residues. This is a way to include an OR statement in Grasp's subsetting vocabulary.

2.11 - Simple Property Mathematics

Sometimes the properties calculated or imported into Grasp are not exactly what one is interested in. For instance one might be interested in not the surface potential interpolated from map one or that from map two, but some average of them (for instance weighted by the average ionization state of two residues). The same might be true of the maps themselves and one might like a combination of the two maps. Simple property mathematics addresses this by providing for arithmetic on one or two fields, putting the result in another or in the same field.

The operations available for maps include addition or division of two maps, along with multiplication by a constant and swapping the two maps. The operations for atom and surface properties include addition, subtraction, division and multiplication of two properties, as well as addition or multiplication of a single property by a constant. Also supported are special functions on a single field including square root, reciprocal, exponentiation, logarithm, cosine, sine, and hyperbolic functions.

More useful to some extent are the contraction operations which take a field and return a single value, maximum, minimum, average and sum. These can be combined with subsetting so that only portions of the selected fields are acted upon. For instance, one can find the accessible area of all lysines, or all charged groups, or all charged groups of a particular helix, etc.

Fields can be shifted around. For instance, one can multiply a field by 1.0 and place the result in a second field. This can be useful if one field is to be used as a dummy field. For instance, if charges were assigned to represent helix forming potential via a DelPhi charge file format they can then be passed on to one of the general property fields.

Properties can be inverted. One can invert the normals of a surface to make it appear similar to its complement. One can also or invert its electrostatic potentials since one might expect a complementary surface to also be complementary in potential. This is simply achieved by selecting the surface of interest and multiplying its surface potential by ­1.0.

Also included is a facility to map an atomic parameter directly to the surface. One advantage of this is that a surface is to some extent simpler than the collection of underlying atoms and as such is often a better vehicle for displaying properties. One further advantage is that by using the accessible surface one can project these properties into space away from the molecule.

2.12 - Objects

An object is an abstraction which represents both the shape and a property of a set of atoms. There are two for proteins and three for DNA.

The first protein object is a backbone trace called a "backbone worm". It consists of a set of cylinders forming a tubular B­spline though atom positions. When the command is made to build a backbone worm, the user is given the default option of using all alpha carbons of the protein backbone or entering a different set. This latter option might be used to select only a subset of all alpha carbons, or to select a totally different set of atoms like amide nitrogens.

If the user has previously constructed one or more worms, the user has the option of replacing all of these or adding to them. There may be up to one hundred disjoint splines constructed.

If two sequential atoms in the selection are more than a certain distance apart the spline is terminated and a new one begun. Hence, if constructing a backbone worm when the protein has more than one chain, there will be one spline per chain. The distance used to determine spline breakage can be set by the user. If set sufficiently large, this will force the construction of a single spline through disjoint chains. It can also be useful in judging patterns of distances of a certain subclass of atoms - spline breakage contains local distance information.

A B­spline segment is constructed from each set of four consecutive atoms in the selection list. Each segment is made of four subsegments by default. This can also be adjusted by the user. Note that this change in resolution only takes effect when a worm is built and does not alter the segment density in chains already constructed. Each subsegment is a tube of polygonal cross­section. The default number of sides to this polygon is ten, and this too may be altered by the user. Finally, the user may alter the radius of the cross section of the worm, i.e. the worm thickness. Changes made to these two parameters are reflected immediately in any worm already built.

One limitation to the subsegment density described above is that the number of subsegments per segment must be even. This arises from the method of assigning atoms to each subsegment. The procedure used is that the first half of the segment is assigned to the second atom in the defining four atom sequence and the second half to the third atom. Hence to be able to equally distribute subsegments there must be an even number of such per segment. Note that this method of subsegment distribution results in no segments assigned to the first and last atoms in a sequence, and only half segments to the second and second-to-last atoms.

The second protein object is a peptide plane representation called a "backbone box". As is well known, the peptide bond has double bond character and so is stiff to torsional rotations. As a consequence the set of backbone atoms CA(n)­C(n)­N(n+1)­CA(n+1) lie in a plane. This can be represented as a quadrilateral with corners at carbon alphas (CA) and at the oxygen and hydrogen of carbon (C) and nitrogen (N) respectively. This is given a little width for display purposes to make a quadrilateral box. These boxes can be colored in several ways. The default color scheme is white at the alpha carbons, red at the oxygen, blue at the hydrogen. In this mode it is easy to see, for instance, where backbone loops have all carbonyls or all amide hydrogens aligned in one direction (which can be significant electrostatically). It is also makes secondary structures particularly clear. Boxes can also be colored by subsetting based on the underlying atoms, or uncolored to display only parts of chains.

The 3 DNA objects are the phosphate backbone, the pentose sugars, and the DNA bases themselves. The backbone can be represented as a thick ribbon smoothly splined through backbone phosphates. The pentose sugars are represented either as rings or line pentagons. The pentagons are color coded by the endo­ or exo­ nature of the sugar carbon. The bases are represented as rectangular slabs colored by base type. Base representation can also be made to width­encode a DNA base pair parameter. Support is provided for the output from the program CURVES which describes 38 such parameters. These can also be mapped to a helical sheet and color coded.

2.13 - Pair-Wise Interaction (Matrix) Representations

One quantity which is difficult to represent by conventional means is pair­wise interactions. This is because the variable has a value and two positions instead of just one position. For this reason this is like attempting to display a matrix, as opposed to a vector. Such variables occur in electrostatics as the interaction energy between each charge in a set of charged sites. Another example is effective inter­residue forces which some have developed to model protein stability. Since both these uses are essentially residue-based, the formulation of the pair­wise interaction representation is residue-based in Grasp - it acts between residues, not atoms.

In Grasp these forces are represented by means of lines running between pairs of sites. These lines have as properties the interaction strength and those properties of both interaction residues themselves. Hence lines may be colored (or "uncolored") by standard subsetting commands.

Interactions also have the property of "rank" - since sites may have several interactions, each interaction also has a rank amongst those assigned to that site. Thus one can subset by rank, and only show the strongest interaction for each site. Since interactions can be strong by being very attractive or very repulsive, the interaction strength very negative or very positive, support is provided for ranking by either criteria. This can be useful in determining 'zones' of interactions - patches of residues which interact mostly amongst themselves.

Grasp goes one step further than merely representing forces with lines by expanding the lines into cylinders. As well as being visually striking, this allows the width of the cylinder to act as an indicator of the absolute strength of interaction. By default, Grasp sets the maximum width to represent the maximum absolute interaction, but this can be set to be larger or smaller by the user. All other coloring operations still apply. Grasp also allows for the cylinder representation without width encoding.

The ends of these lines or cylinders are by default set to the average position of all atoms in the residue. This can be altered to any subset within the residue. For instance, in some cases the position of the residue charge might be more appropriate, in others the alpha carbon, or center of the side chain, etc.

Since interactions are be distance dependent, the user can explore this dependence by multiplying or dividing each value by the distance between its two sites. This scaling can help the user determine which interactions are unusually strong or weak.

Matrix values are not calculated at by Grasp. They must be imported via a data file, the format of which is described in appendix A.

2.14 - Stereo/Split Screen Operations

Stereo viewing has traditionally been achieved by duplicating the view, separating the views by a certain distance so that there is no overlap, and then giving the right­most view a twist of about 8 degrees about the vertical direction. Grasp follows this approach and extends it to a 'split­screen' capability.

The stereo separation and twist are under user control (tests within our labs showed conclusively that everyone has their own preferential stereo twist). Separation and twist are under mouse and dial control and twist may also be entered explicitly.

The duplicate view can be treated as completely independent, so that nearly all display possibilities can be used either on the left or the right. This allows the user to display alternate views side by side. For instance, one might want to view the surface color coded by potential and also by curvature at the same time. The left and right views can also be superimposed. One can also manipulate either view independently. For instance, one can display the front and back of a molecule simultaneously. Formal subsets in each view are also independent, so different arrangements of such subsets can be portrayed in right and left views.

3 - Getting Started

3.1 - Grasp Environment

There are a few things one should check before running Grasp.

First is to ensure that one has write privileges in the directory the command is issued from, which can often be a problem if working from someone else's directory. Grasp needs this permission to enable it to write temporary files, which are removed upon exiting the program, and some permanent data files, such as a color map if the user alters those provided, and also "error" files if it detects odd situations (such as finding too many bonds for an atom when reading a pdb file, or fractionally charged residues upon reading a charge control file).

Second, Grasp reads in a few data files upon startup. It needs to know what directory these files are in. To do this, it reads the environment variable GRASP (note capitals) which should be set to the directory with all the files with the extensions ".dat" or ".crg" or ".siz" or ".gs". For those not familiar with Unix, the command to do this is

setenv GRASP dirnam

where dirnam is the directory name. One should place this command in the file .login in one's home directory so it is read and executed when the user logs in. One can check the value of this variable by entering,

echo $GRASP

Grasp also has a directory of "last resort" if it can not find the directory defined by $GRASP. The backup directory it looks for is ./aakdat, i.e. in a directory one lower than the user is in called aakdat. These data files are listed in the appendix and involve such things as default radii for atoms, default charge sets, and information used in surfacing molecules.

Third, Grasp will check for a file called .init_Grasp. This file can contain commands which set variables within Grasp, such as default display modes. These commands are listed in Appendix B. Grasp searches for this file in three places. First it checks the directory defined by $GRASP, second it checks the user's home directory, and last it checks the local directory from whence the command was issued to start the program. The purpose of having it check all three locations is to allow for hierarchical control of Grasp settings. For instance, one might want to set some parameters for all users, in which case they are set in the $GRASP directory. Individual users might want different parameters for their own work, and so alter the file in their home directory. Finally, the individual user might find that for some projects different parameters are better - small molecules might want one set of display parameters, large molecules others, in which case control should be via the file in each particular directory. The order the files are read is important because if two files set the same parameter, preference is given to the later file.

3.2 - Running Grasp

Now you can start Grasp simply by typing "grasp". Once this command is issued, and the appropriate files searched for and read, the default graphics window opens and shows a set of axes or cross­hairs in the X, Y and Z directions. The Z direction is towards the viewer with positive nearer and negative farther away, the X direction is left to right, with right positive and left negative, and the Y direction is up and down with up positive and down negative. The cross hairs run between +/-1 in Grasp internal coordinates. To help visualize this domain one can view the Grasp box. This is done by pressing Control O.

3.3 - Dial and Mouse Movement

One moves the view by using either a dial box or the mouse. The dials work accordingly:

The mouse moves the view by depressing the left or middle buttons or both and moving the mouse:

Left button:
Rotations about the axis perpendicular to the direction of mouse motion.Hence there is a sense of "rolling" the molecule as if the molecule where resting on a solid surface in the XY plane and the cursor was the user finger.

Middle Button:
Up and down moves the molecule away and towards the viewer respectively. Note that this is NOT a scaling, but an actual motion in the Z­direction and hence corresponds to pulling the molecule towards or pushing it away from the user. Left or right motion rotates the molecule about the Z axis.

Left and Middle Buttons Together:
Translates the molecule in the XY plane in the direction of mouse movement.

This implementation of dials via the mouse (mouse­dials) is a little different from some programs since only two mouse buttons are used. This is partly because only two are actually required to allow the six independent rotations and translations, but more importantly because the third button, the right button, is reserved exclusively for the menu interface.

One further convention adhered to in Grasp is that where appropriate, the middle button adds and the left button subtracts. For instance when adjusting the indexed colors in Grasp the middle button increases a color component, the left button decreases it.

The rate of rotation or translation, the sensitivity to mouse or dials, can be set via the menus. It is also possible to assign different functions to the mouse, such as surface scribing, or projection plane position. These are accessed via menus.

Note that the box does not rotate with the the cross hairs. The significance of this is that box represents a space which is invariant with respect to the user. Rotations and translations do not actually affect the molecule coordinates, only the viewing of them. One way to think of this is that it is not the molecule which is moving but actually the user and the box (since both move the box appears stationary). The language used in Grasp is of "original" coordinates, which "belong" to the molecule, and "box" coordinates which belong to the user (and hence the box). One can change molecules into the users frame of reference, i.e. make rotations "real", via formal subsetting and some options for file export.

3.4 - Data Files

One can now read in one of three type of data files, atom files, surface files or maps (3­D grids). Note that instead of reading in a file once the program has executed one can give the name of a data file:

grasp lys.pdb

which will load in coordinates from the pdb file lys.pdb. Other than pdb files one can also give the names of maps, with the extension ".phi", or surface files with ".srf". If the name of the file does not have one of these three extension then the program will prompt the user as to which type of file it is.

Reading a data file from within Grasp involves using the menu system. Clicking the right mouse button, then clicking on Read and then on one of the primary data file types will prompt the user to enter the file name, select a default file, or see a list of files. The list will be all files in the initial directory which have the correct extension for that file type.

3.5 - The Textport

When character input is requested in Grasp, it is via what is called the textport, which is the character based window from which Grasp is initially launched. To enter information to the textport the cursor has to be positioned over this window. If Grasp is expecting information, e.g. expecting a file name, it tries to make this easy for the user both by automatically positioning the textport over all other windows, and by automatically placing the cursor over this window. And when the information input is complete and return has been hit, the cursor will automatically jump back to the spot on the graphics window it was before the request for information. Similarly if the user wants to type a command, the cursor will automatically place itself over the textport when the user begins to type, jumping back when return is hit.

Note that in both these examples the cursor starts over the graphics window and ends there too. The user should NOT attempt to move the cursor onto the textport except in the following two cases. Sometimes the user may have moved the textport, or resized it, and as a consequence the cursor may miss it when made to "jump" by the program. Also possible is that the cursor will come to rest upon the textport when instructions are not being entered. This causes a "change in input focus" for the program i.e. it expects input from the textport rather than from the graphics window. When this happens, for instance, the molecule will not rotate when the dials are twiddled because the program is not "listening". This question of "focus" can often give beginners the most problems in getting started with Grasp, so when in doubt check the cursor position.

Hitting return when the cursor is over the graphics window causes the textport to alternate between background and foreground, i.e. being behind all other windows or on top of them. If the user has resized the textport, for instance made it bigger to review more information, or repositioned it, hitting return will also resize and reposition the textport. Another use of this is that pushing the textport and then bringing it back will usually force a redraw. If for any reason the graphics look funny, for instance 'damaged' by the movement of some other window or some other program, or if the initial view upon starting the program looks strange, this is a simple way to redraw the view.

If one has read in a pdb file or a srf file there should now be something displayed from within Grasp, either a molecule or a surface. The default display of either of these can be altered as described later. Upon reading a structure (i.e. atoms or surfaces) a scale is assigned to the unit box, i.e. the width in Angstroms is calculated (and written to the textport). This is calculated such that the structure will fill up two thirds of the the box in its longest dimension. If the user instead has initially read a potential map the scale is such that the potential map will fill the unit box exactly, i.e. the boundaries of the potential map are at +/­1 in each direction. This scale is now set for the duration of the Grasp session as there is as yet no facility for altering global scaling. The view can now be manipulated, quantities calculated, structures built, etc.

3.6 - Exit

Exiting the program can be done three ways. The first is NOT recommended except in emergencies (the program has inexplicably locked up) and that is to put the cursor over the textport and hit Control C. The normal way to exit is either through the main menu or via Control Q. If the program is correctly exited one should notice that the cursor is no longer yellow, which it is during normal Grasp operation, unless of course this is the user's normal color for the cursor.

4 - Menus

Menus carry most of the functionality of Grasp. All menus are accessed via the right mouse button. All selections within a menu must also be chosen with the right mouse button. Menus appear when this button is depressed and remain when the button is released. Note this first release of the right mouse button does NOT select an item. The program is essentially frozen until the right mouse button is depressed and released AGAIN.

If the cursor lies over a menu entry when the button is released the second time, then that menu item is chosen. If the user "clicks away", i.e. releases the button when the cursor does not lie on a menu entry, function will abort or in some cases will continue with a default value. For instance, when altering the molecular surface, first the user will get a submenu for the quantity displayed on the surface (potential, distance, color, etc.) and then a submenu for the draw mode (mesh, lit, points, etc.) of the surface. If one does not want to change the quantity displayed, one clicks away from the quantitiy menu and continues on to the draw mode menu. On the other hand, if the user chooses the Build entry in the main menu and then clicks away the program exits from the menu altogether.

Some parts of some widgets, such as the color scale, are sensitive to the right mouse button. This means if the right mouse button is depressed while on this area, the normal main menu will not appear and instead the user will get a menu associated with that widget. Clicking anywhere else in the graphics window will bring up the main menu.

For ease of description, this document will use the following notation to describe a sequence of menu operations: "Display: Stereo/Split Screen: Dials to Both" means choose Display from the main menu, then Stereo/Split Screen from the Display menu, then Dials to Both from the Stereo/Split Screen menu.

The menu options will be described in the order they appear in the main menu, starting with Display. Submenus will be described where they apppear.

4.1 - Display Menu




Hide ALL

Stereo/Split On

Show displays a structure, Alter changes the display of a structure, and Hide causes a structure to disappear from view (this is not the same as causing a structure to disappear by coloring it 0 - when a structure is hidden in this manner, it retains all its characteristics including color). All of these options produce a submenu which allows the user to choose from the following structures: Molecular Surface, Atoms, Bonds, Cavities, Objects, Contours, Vectors, and Interaction Matrix.

For Molecular Surface, the user may choose from several coloring schemes: potential, curvature, discrete colors, distances, general property #1, and general property #2. Surface draw modes are rendered, rendered but not lit (pseudo-lit), mesh no rendering, points, transparent, solid/mesh mix, and solid/trans. mix.

For Atoms, the color options are atom type, charge, potential, distances, property #1, and property #2. Atom draw modes are flat circles, full spheres (CPK), flat patterns, little bond-atoms, line spheres, and point spheres.

For Bonds, the coloring choices are user defined, which lets the user set them (default color=1), atom colors, which adopts the underlying atom colors, and saved set, which applies an internal color scheme. Bond draw modes are lines, sticks, and rods. Rods are very slow to draw.

For Cavities, one has the option of coloring as the molecular surfaces, coloring by number, or coloring by number patches.

For Objects, the user must first select the type of object: backbone worm, backbone boxes, DNA bases, DNA backbone, DNA sugars, DNA axis, ellipsoids, DNA H-bonds, or distance line. DNA objects come with several possible options. Most of these are only appropriate if data in the form of a Curves file has been read in, because the options refer to which of the many Curves parameters are to be displayed.

For Contours, colors as well as values are set when contours are built. Contour draw modes are the same as surface draw modes. Control of depth cueing for contours is controlled by "Set Parameters: Depth Cueing".

For Vectors, the possible types are electric dipole, which is a large 3­D arrow of length 0.3 box units, electric field vectors, which can be drawn of constant length or with length dependent on the field strength, and electric field lines, which can be drawn as lines or tubes. The user selects a color for the molecular dipole vector when calculated. For field vectors the user can specify the maximum vector length and the field strength this should correspond to, but only when these quantities have been calculated. Note that it is necessary to calculate a vector quantity before displaying it. The cylinder mode for field lines can be very slow to draw.

For Interaction Matrix, the choices are lines, cylinders, width-encoded cylinders, and extras. The width of the width­encoded cylinders depends upon the absolute value of the interaction. This value is divided by a "maximum strength" value which can be changed, rounded off to unity if greater than one, and multiplied by an internal width to give the resultant cylinder thickness.

The extras option presents a submenu with several entries. One is to refine residue centers, which alters the exact coordinates of the point within each residue each strand emanates from. Upon choosing this option, the user is prompted for a selection on the command line. The user could then input the command "a=ca" whereupon the strands would begin on each residue's alpha carbon. If more than one atom is selected for a residue, the strand begins at the average position of those atoms. Another option is to set the maximum interaction strength for width­encoding as described above. Also included are options to multiply or divide the interactions by the distance between interaction sites. The new maximum and minimum values are written to the textport upon each use of these options. Note that if the draw mode is set to variable widths then this can affect the relative widths of the strand cylinders. To maintain a similar spread of widths the following process of rescaling the maximum cylinder width value is enacted. The ratio of the largest (absolute) interaction value (before distance scaling) to the value set for the maximum width is found. Then the largest (absolute) value is found after distance scaling, and the value for maximum cylinder width set so the ratio just calculated is maintained.

Hide ALL clears the view completely.

The Stereo/Split On controls the stereo display options. It produces a submenu with entries Dials to Both, Dials to Right, Dials to Left, Right­Hand Twist, Stereo Parameters, and Stereo/Split Off. This menu can be accessed directly with Control S. The first three entries decide which side is going to be "attached" to the rotations and translations as entered by the mouse or dial box. The default entry for this menu is Dials to Both - both views are moved equally. In this mode, the two views differ only by an imposed separation of 0.5 box units in the X direction, although the views may not appear identical due to the perspective automatically included in all Grasp views. Choosing Dials to Right or Dials to Left allows one view to be spatially manipulated independently of the other. Note that if the user has defined one or more formal subsets, only the views of the subsets on the side for which the dials are attached can be moved.

Right­Hand Twist causes the right hand view to be twisted in an axis running through the center of its world coordinate system in the Y direction (vertical) by a certain angle This twist is set to 8 degrees by default. This value can be altered in several ways. If the user has a dial box, the left bottom dial will alter the stereo twist. The user can also fix the Z­rotation function to stereo twist via "Mouse Functions: Alternative Z­Trans. Alternative".

Stereo Parameters presents a submenu to allow the user to restore twist to the default value, enter a twist value, enter a separation value, or remove twist and stereo separation altogether and superimpose the left and right views.

Even if the user is in "twist" mode the user can still independently manipulate the two views spatially. This is not recommended since the purpose of twist mode is to allow stereo viewing which requires the two views to be essentially identical except for the vertical twist.

If while in split screen mode, the user attempts to change display properties, Grasp will prompt the user on which "side" the changes should be made. For instance, if the user attempts to hide the bonding display the choice is left, right or both. Thus the display on the left can be representing one facet of a molecule (such as electrostatic potential on a surface) while the right represents another (such as atomic B­value).

Stereo/Split Off quits stereo/split screen mode. The left­hand orientations (world and subset) are the ones retained for the single­screen view. All differential display characteristics the user may have applied to the right view are lost. However, the stereo twist value is still stored and may be retained throughout a session.

4.2 - Build Menu

Molecular Surface

Accessible Surface

Backbone Worm

Backbone Boxes


DNA Boxes


Consensus Volume

Ellipsoidal Objects

The build and calculate menu options are easy to confuse. For instance, does one build a contour or calculate a contour? Are field lines built or calculated? In general, build deals with calculating the data intrinsic to a display structure, such as a surface or backbone representation or internal cavities, whereas calculate provides numbers which may or may not be related to such structures, such as potential maps or volumes of surfaces.

A Molecular Surface or an Accessible Surface is made by essentially the same algorithm. When choosing whether to construct one or the other, the user has to consider two options. The first is which atoms to use in forming the surface, and the second is whether to add this surface to previously constructed surfaces, or to overwrite them. Note that this is the only way to delete a surface within Grasp. The menu for selecting a subset of atoms is: All Atoms, A Molecule, A Format Subset, and Enter String. All Atoms selects all stoms, A Molecule presents a menu containing all molecule numbers, A Formal Subset presents a menu with the names of all atom­based subsets, and Enter String will cause the cursor to jump to the textport and wait for the user to enter a subsetting command. If no atoms are chosen at the end of this procedure, the routine aborts.

The process of constructing the molecular surface occurs via the construction of a temporary accessible surface. A correspondence between the vertices of this intermediate surface and the underlying atoms improves the accuracy of the final surface. It also allows for a unique mapping between atoms and molecular surface such that each accessible surface vertex is assigned an underlying atom, and each molecular surface point is assigned an accessible surface point. The combination of these two assignments leads to the association of each molecular surface point with an atom, an association which is termed "contact" within the program.

The process of surface formation will cause plenty of information to be written to the screen, including the scale at which the surface is constructed and the number of vertices and triangles in the completed product. The information here can be useful in debugging (for example the total number of vertices might exceed the maximum allowed number).

Surfaces should appear automatically after being calculated. They will not be colored however. This must be done by the user via whatever method chosen, by calculating potentials at the surface, calculating curvature, etc.

The Backbone Worm and Backbone Boxes are built for all current molecules. The only options associated with these objects are for the backbone worm, which allows the worm to be built for all CA atoms or a subset. If these objects fail to appear after construction, go through the Display menu explicitly. The backbone worm can be slow to display. Sometimes this can be put to good use. For instance, if the user switches to single buffer mode (Control R) while the worm is being drawn, the path of the chain from N terminus to C terminus is nicely illustrated. The backbone worm only requires the carbon alpha positions to be correctly produced, while the backbone boxes require also the carbonyl oxygen and amide nitrogen positions to successfully complete construction.

Building Cavities/Connectivity causes the program to check all vertices for connectivity. The first point, or seed point, is chosen at an extrema, and so can not belong to a cavity. All points associated with it, those which can be reached by travelling along triangle edges, are deemed the "non­cavity surface". All others belong to cavities. Note that this will give an incorrect assessment if there is more than one disconnected constructed convex surface. For this reason the user should calculate cavities on the surface of the whole molecule, not subsets. Printed to the screen are the number of triangles that make up each cavity found. Cavities are automatically displayed in the same display mode as the molecular surface. They can be sequentially colored.

Building DNA Boxes requires that a DNA PDB file has been read in. (Note that mixed files containing DNA and protein are fine.) Display should be automatic. Grasp can handle structures with up to four independent backbone strands.

Contours can be made in 3-D, 2-D with an interpolation plane, or 2-D with a molecular surface. These all require two inputs by the user: the isopotential value, and the color to be assigned to that contour. The latter should be a color index, an integer between 1 and 99. One can enter more than one isopotential value to create more than one contour at a time, as long as one enters the same number of colors. For instance:

enter 3-D contour value

>> 1.0,2.0,3.0

enter 3-D contour color(s)


will create contours at one, two and three kt, and give them colors 1, 2, and 3 (red, green, and blue). To delete a contour (which may be necessary to make room for new ones), one goes through the same procedure as making a contour of the same isopotential value, one gives it color 0. Note that this remove is NOT the same as hide or uncolor, as it actually removes the contour data from the program.

Although the usual use of building contours will be isopotential contours, it can be used for more varied purposes, since the actual contents of the map are irrelevant to the contour facility. For example, one can contour a DelPhi "eps" map if one has read one in, or one can contour a consensus volume map (see below) if one is calculated. One should remember that there are two internal maps in Grasp. The contouring proceeds on whichever map is "current". This is usually map 1, but will be map 2 if one has just read in a map. Set the current map with "Miscellaneous: Change Current Map".

Consensus Volume produces a map - a 3D lattice of values. It is calculated by adding the value 1.0 to each grid point which lies within the Van der Waals volume of each molecule. Thus if there are five molecules and a grid point lies within all five, it will be assigned a value of 5.0. Some points will fall within only some molecules. This map can then be contoured at any level desired. For instance contouring at the level of the number of molecules will give the surface of the volume common to all molecules. At present there are no options to this facility and all atoms are included in the calculation. The map is stored as map 2.

Ellipsoidal Objects constructs ellipsoids enclosing the Van der Waals surface of the atoms, residues, or surfaces selected. This selection can be all residues, an atom subset with one ellipsoid, an atom subset with many ellipsoids, a surface subset with one ellipsoid, and a surface subset with many ellipsoids. For the atom options, the user is given the choice of all atoms, a molecule, a formal subset, or entering a subsetting string. For surfaces, the choices are all surfaces, a single constructed surface, a surface formal subset, the currently scribed surface, or entering a subsetting string. If ellipsoids have already been constructed, the user has the option of adding to the current set or replacing them.

4.3 - Calculate Menu

New Potential Map

Pot. via Map at Surfaces/Atoms

Surface Curvature (+Display)

Simple Property Math

Dipole Moment

Field Lines

Field Vectors

Volume of a Surface/Molecule

Area of a Surface/Molecule

Distance Array

H-Bonds (DNA Bases Only)

The calculate option does much that is unique in Grasp. It allows the user to quickly calculate electrostatic quantities like maps, fields, and site potentials, as well as curvatures, distances and volumes, and manipulate fields of information previously calculated or imported to the program.

Calculating a New Potential Map causes the program to execute its internal Poisson­Boltzmann solver. The size (in Angstroms) of the map produced is automatically determined. Only the linearized Poisson­Boltzmann equation is solved. Such parameters as the probe radius, ion exclusion radius, salt concentration and inner and outer dielectric values can all be set via "Set Parameter :Electrostatic Parameters". The map produced is stored in internal map 1. If no charges are assigned, then the procedure will inform the user and abort rather than calculate a null map.

It is important to realize that by default all atoms and therefore all charges are used in the calculation. If the user wants to perform a calculation on a subset of atoms, those atoms not required must have their radii and charges set to 0. The algorithm will ignore any atoms with radius 0 so that they will not contribute to the water exclusion (=low dielectric) volume, however, it will NOT ignore the charges on these atoms. Therefore one needs to neutralize the charges on the unwanted atoms as well. Since changing radii to 0 also causes atoms not to be displayed, the user ought to remember to reset those atoms to their correct radius after the calculation, for instance by reading in a default size file.

A typical use of the above procedure for removing some atoms from an electrostatics calculation is one where the original crystallographic coordinates are included for several water molecules. It is always an interesting question as to whether such waters should be treated as low dielectric, and constrained in their motion, or high dielectric, and bulk water. A good rule to use is that if a water is highly coordinated to the protein, so that it makes two or more hydrogen bonds, a case can be made for low dielectric, otherwise set it as high by setting its radius to 0 in the calculation and turning off any assigned charges.

As with building surfaces, much information is written to the screen during the calculation, some of which is useful to the user in verifying the accuracy of the calculation, that it has converged, that the correct total charge has been assigned, that the scale of the final map is approximately correct, etc. A typical calculation should take about five seconds on a Personal Iris.

Pot. via Map at Surfaces/Atoms calculates the potential at all surface vertices and all atoms from the current map. If one has just calculated a new potential map, this is map 1. The algorithm uses trilinear interpolation from the eight grid points which make up the map grid cube which an atom center or surface vertex falls within. If it lies outside the map, a 0 value is assigned. Note that this process will overwrite any previous potentials. If this is a problem, the user should first store the previous array in one of the other variables, as described under Simple Property Math.

Surface Curvature (+Display) will cause the program to calculate the curvature, as defined in Nicholls et al., for a set of surface points and a set of atoms. As our definition of curvature is related to accessibility of water to a single water placed in contact with a particular surface point, the set of atoms chosen is crucial, since the hard sphere radii of these atoms will determine this accessibility (note that a 0 radius atom does not affect accessibility). Thus, for instance, if one wants to compare the curvature of two surfaces which make up an interface, one should choose surface 1 and the atoms which made surface 1 for the first calculation of curvature, then choose surface 2 and its atoms for a second calculation.

Curvature calculations can sometimes take a long time. This is due to the choice of test sphere used to determine accessibility - more points take more time. The choice of this density is made automatically relative to the scale employed in the surface creation. Because Grasp has only a fixed number of test densities, the time taken in calculation will vary considerably.

Upon completion of a curvature calculation, the display should automatically switch to displaying this quantity. The values are scaled to +/­ 100, such that 100 would imply that the surface point is completely accessible, which will never happen since this would imply that one could put a water molecule anywhere in contact with the original water molecule touching the surface. However ­100 implies that the surface water is completely isolated from other water molecules, which is quite possible, for instance in a deep cleft or if the surface is enclosed inside the molecule or is part of a molecular cavity.

Simple Property Math is one the most useful options in Grasp. Its submenu gives the user these choices: Potential Maps, Atom Properties, Surface Properties, and Map Atom Value to Surface, and Map Atom Value to Worm.

Potential Maps allows the user to perform an operation (add, subtract, multiply, divide) between maps and store the result in either map , to add or multiply by a scalar value, to perform a function on a map (square root, reciprocal, raise to a power, exponentiate, absolute value, natural log, sine, cosine, hyperbolic cosine, hyperbolic sine), or to swap maps (put map 1 into map 2 and map 2 into map 1).

One of the scalar map options is to apply a "convex" correction. This was added after it was noticed that potential maps generated on the the Convex used for most of our DelPhi calculations where misread if transferred onto an Iris. To be more precise, all real numbers (but not integers) were exactly four times too large. This includes the potential map center and scale as well as the potential values. This option then will rescale all those values to their correct value. In addition, if the Grasp box scale has been derived from a potential map, then the user is prompted as whether to reduce the box scale by one quarter as well.

One should note in "map math" that no checking is done to see if the grids actually have the same center and scale, i.e. whether they refer to the same part of physical space. The values of corresponding grid points are just added, multiplied, etc. The inclusion of a division operation was prompted by the desire to calculate effective dielectrics, the ratio of the potential calculated with Coulombs Law to that via the Poisson­Boltzmann equation. If 0 is found in the denominator of a division, then the new grid value is set to 0.

Math on Atom Properties and Surface Properties are the same in that the same basic operations can be applied to either. These operations are add, subtract, multiply, divide, multiply by a constant, add a constant, maximum, minimum, average, sum, and special functions (square root, reciprocal, raise to a power, exponentiation, absolute value, natural log, sine, cosine, hyperbolic sine and hyperbolic cosine).

Add, subtract, multiply, and divide require selecting three properties (two properties for calculation and a third to store the result). Multiplying by a constant, adding a constant, and special functions require two selections (the property acted upon and storage). Maximum, minimum, average, and sum return a single value printed to the textport. Maximum and minimum also print the atom or vertex number with with the value is associated.

Properties for each step of the math calculation are selected from a property list for either atoms or surfaces. The user is also prompted for a subset of all atoms or vertices to do the math upon, which is then common to each array. The user should take some care in using this simple math facility to ensure that the property arrays are entered in the correct sequence.

Map Atom Values to Surface requires that the user select a surface, a set of atoms, an atom property, and surface property. The connection between surface and atoms is via the original construction process, i.e. which atom is responsible for which part of the surface. If the responsible atom for a particular vertex in the surface selection is not included in the selection of atoms made, then no value is set in the property array for that vertex. Most typically the user will apply an atomic property universally, i.e. choose all atoms and all vertices, to the surface.

Map Atom Values to Worm requires that the user select a surface, a set of atoms, an atom property, and surface property. The connection between surface and atoms is via the original construction process, i.e. which atom is responsible for which part of the surface. If the responsible atom for a particular vertex in the surface selection is not included in the selection of atoms made, then no value is set in the property array for that vertex. Most typically the user will apply an atomic property universally, i.e. choose all atoms and all vertices, to the surface.

The flexibility of operations on atoms and vertices greatly exceeds that of maps, which is mainly because the uses so far imagined for the former greatly exceed those for the latter. In time it is hoped that Grasp will support a math interpreter rather than use the menu system which should more fully realize the potential for on-the-fly analysis it is hoped this function makes possible.

Dipole Moment takes all the charges on all atoms and calculates a dipole and, if necessary, monopole. The latter is necessary if the total charge is not zero. The procedure is to find the sum and charge weighted average position of all positive charges and the similar quantities for the negative charges. If the two sums are not the same, then there is a monopole equal to the sum of the sums acting at the charge weighted average position of the larger (in absolute terms) of the sums, plus a dipole of magnitude determined by the smaller (in absolute terms) sum multiplied by the distance between charge­weighted centers. These values are printed to the screen, plus once calculated the user can display a dipole "arrow" centered at the average of the charge weighted centers, and of length 0.3 box units. This is displayed via "Display: Show: Vectors".

There is as yet no option to display or calculate multiple dipole vectors.

Field Lines are a difficult quantity to visualize. This is mainly because they are infinite in number and it is difficult to visualize an infinite number of lines. In a "correct" implementation the number of lines within a given volume is proportional to the average field strength. Hence one could imagine discretizing this concept so that there are a finite number of such lines. However this is NOT the approach followed in Grasp. Instead one chooses a set of starting points, or seed points, and projects the direction a positive or negative charge would take from each such point, the field direction, as found from the current potential map. A line segment is drawn a certain distance in this direction and then the field at the end of this segment calculated. This process is repeated up to a fixed number of times to give a field "line". Typically one hundred segments are calculated. If the length of each segment is small enough, the line appears smooth.

Seed points are determined by the first submenu of this selection. The choices are: Center of Atom Set, Surface, Atom Set, Origin, and Enter Coordinate. Center of Atom Set will prompt the user for a set of atoms. The average of these coordinates will then be found. If the number of lines chosen in a later menu is greater than one, then that number of random positions within half an Angstrom of this position are found and used as seed positions for field lines.

In selecting Surface, one can obviously choose all surface vertices, but often a more useful selection is some patch of surface selected by surface scribing. Once chosen, a number of points equal to the number of lines chosen are found at random over the selected surface as seed points.

Choosing Atom Set will cause the center of each atom selected to be initial seed points. If more than one field line is selected, that number of seeds will be formed within 0.5 Angstroms of each atom center. The user should note that there is a maximum of 100 field lines and should be careful not to exceed this limit or the process will not run to completion.

Choosing Origin causes the initial seed point to be at the center of the Grasp box. This number of seed points can be multiplied in the same random manner as described above.

Finally, Enter Coordinates allows one to enter the absolute coordinates for the initial seed. Again this can be multiplied by random scatter about this point.

The next menu deals with field line directionality. Clearly one has a choice of whether to use a positive or a negative "test" charge when plotting the field line. Traditional electrostatic lines begin on positive charges and end on negative charges (or infinity). In Grasp they can go either way depending on the sign of the test charge, or go both ways! Note that by choosing bidirectional lines one actually creates two field lines, one in each direction. If one has chosen to create the field line seeds from a surface then one has yet another choice, whether one would like to force the direction of the field line into or away from the molecular surface.

Next one can choose the number, or multiplicity, of field lines. As mentioned above, this will multiply the initial seed positions by finding random positions in the vicinity of the initial seed point(s). The default value is one. As mentioned above, bidirectional lines count as two. Thus if one chooses the atom centers of 20 atoms, with a multiplicity of 2 and choose bidirectional lines one actually gets 80 field lines.

Finally, the user gets a choice of colors for the field lines, the default being white. If one has previously created some field lines, then one has the choice of adding to those already created, or replacing them (Note the 100 field line limit applies to the total number of field lines). Field lines can be drawn as lines or as cylinders. The latter display is more dramatic than the former but is very slow to draw.

Currently in Grasp the line segment length of the field line is constant. This means that sometimes the lines will exit the potential map i.e. for which there is no interpolated field. Lines will then terminate. Another problem is that around charges field lines will not actually terminate, rather they oscillate. This is due to the nature of the field around a charge mapped onto a grid. There is also no method of illustrating the field strength at any point along a field line. These shortcomings will be addressed in future releases.

Field vectors can be calculated at up to 100 sites. Sites of interest are chosen as either a set of atoms (each atom center), the center of a set of atoms, at a random selection of points from a surface, or from a point whose coordinates are entered by the user. Once the points of interest are selected, the magnitudes and directions are calculated from the current potential map. Each result is displayed as a field arrow similar to that for the molecular dipole, originating from each chosen point.

The user is given the option of giving each field vector either a constant length, which is then entered in Angstroms by the user, or a variable length, where the user enters both the maximum vector length in Angstroms and the maximum field strength to which this corresponds. In the latter case field vectors of strength less than the maximum representable field strength are assigned a proportionally smaller vector and those greater than the maximum the maximum length. Note that these lengths can not be altered once set and neither can the color for which the user is prompted for each set of vectors. As with field lines, one can delete current vectors when making new ones or one can add them, with the same maximum of 100 total vectors.

One should note that, as with field lines near charges, field vectors may be less than reliable. One should bear in mind the scale of the grid used in calculating the potential map. Fields are calculated by first estimating the fields at the eight corners of the grid cube that encloses the chosen point and then using trilinear interpolation on that set of vectors. The relative error of the field when a point is near a charge is greater than that of the potential because it involves quantities closer to the charge (i.e. higher order derivatives). For this reason one should be very careful in interpreting field strengths quantitatively from any such interpolation when near atomic charges.

The Volume of a Surface/Molecule is relatively simple to calculate, given a tessellation of the surface. One merely chooses a point, preferably close to the surface or molecule, and then calculates the volume of each pyramid formed by this point and the three points of each triangle in the tessellation, being careful to have the order of the points in the triangle base consistent with the surface normal. One can calculate the volume of any closed surface, such as a contour, a cavity, a molecular surface, or an accessible surface. The volume of a set of atoms may also be calculated this way, by tessellating the Van der Waals surface of that set of atoms and calculating the volume of that closed surface. Alternatively, one can map the atoms onto a fine grid, find which grid points are outside and which inside, and form a lattice approximation that way. Since both are simple, Grasp does both and answers are written to the textport.

An Area of a Surface/Molecule is also simple given a tessellation, since it is just the sum of the areas of all triangles. The area may be of any subset of surface one can categorize. The area of a set of atoms is calculated using the surface area algorithm of Grasp which is an efficient version of "Shrake and Rupley", which places points on a test sphere about each atom and finds which are inside and which outside. The density of points, and hence the accuracy used in the calculation, can be altered by the user via "Set Parameters: System Miscellaneous: Surface Area Probe Density". The user also has the choice of whether to calculate the accessible area, as if the radius of each atom is increased by the diameter of a probe, or the Van der Waals surface, with probe radius 0. This probe radius may be set by the user via "Set Parameters: Probe Radii Values", the default being 1.4 Angstroms.

When a subset of atoms is used, the area is calculated as if other atoms do not exist. As with curvature calculations, other atoms are not used to determine accessibility. All results from the calculation are placed in the accessible area array, which can be manipulated via the simple math options. Hence if one wants to know the total accessible area of all lysines in a protein one does not want to select just lysines, rather choose all atoms, then go through "simple math" to calculate the sum of the accessible areas of that subset of atoms. On the other hand one might want to know the total accessible area of a subunit of a quaternary molecule if it were isolated from the total structure. Then it would be appropriate to select just this set of atoms.

A Distance Array can be calculated for any subset of atoms or vertices to any other set of atoms or vertices. It represents the minimum distance for a "from" set to a "to" set. Since atoms have radii, it is often useful to take account of this in the "distance" quantity and subtract it so that the distance calculated is the distance to or from the Van der Waals surface of the atom. Note that this can result in negative distances.

The results of the calculation enter the atom or surface array for distance only for the "from" subset selected, not for both. For instance, if one wishes to calculate the distance map between two surfaces, one should choose this function for the first surface against the second and then repeat it for the second surface against the first. Note the user should be careful that the second set does not contain some of the same points as the first set or the minimum distance will clearly be 0 for these points (or negative if Van der Waals radii are being subtracted).

Distance calculations are quite slow if many points are being checked against many points. At this stage of the program, these calculations have not been optimized. A rough estimate of the time taken for a calculation to complete is to multiply the number of points in each set (the "to" and "from" sets), divide by one million and multiply by two. This is then an estimate in seconds of CPU time. If one is calculating for a 10,000 vertex surface to a 15,000 vertex surface, sit back and wait!

Note: if one wishes to remove the Van der Waals radius from the "to" set, the set of atoms not having distance calculated for, then strictly each distance comparison should involve a square root. Because this is very slow on the Iris, and because this is already a slow calculation, Grasp actually calculates the minimum center-to-center distance first and then subtracts the Van der Waals radius of the atom found closest. This is not strictly correct and may result in inaccuracy at small distances (i.e. of the order of a few radii). This limitation will be removed when more efficient distance algorithms are installed.

Calculating H-Bonds (DNA Bases Only) requires that a DNA PDB file has been read in.

4.4 - Mouse Functions Menu

Atom Information

Surface Information



Command Line Mouse

Z­Trans. Alternatives

There are three different ways information can be sent from the mouse to the program, namely holding a button down and releasing it without moving the mouse ("picking"), which usually returns information on the underlying object, holding a button or buttons, down and moving the mouse, which typically causes rotations or translations, and moving the mouse without any buttons depressed, which usually in Grasp has no effect. These methods can be affected with the menu options within this menu.

Atom Information and Surface Information allow the user to choose whether the left or the middle button is associated with returning surface information or atom information when either an atom or vertex is "picked". The default arrangement is that the left button gives atom information, the middle button returns surface information.

Measure allows the user to associate certain measurement functions with the picking mode of either the left or right buttons which are set as described above. The measures for atoms are InterAtom Distances, Interbond Angles, and Interbond Torsions. These measurements require two, three and four atoms respectively as data. As an example, suppose the user links InterAtom Distances to the left button. The user then "picks" an atom. The next atom picked by this button will cause the distance to be calculated between the first atom and this atom, and the result written to the textport. The next atom chosen after that will cause the distance between it and the second atom to be calculated and presented. This "chaining" of distance calculations continues until the user either double-clicks on the last atom or "clicks away" (clicks on empty screen). This resets the "memory" of atoms so that the next atom picked does not cause a distance calculation but acts as the first atom chosen for a new chain of distances. The same procedure is followed for Interbond Angles and Interbond Torsions, except that the angle calculation occurs for the last three atoms picked, and torsion for the last four. Clearing the "memory" is done as before. Note that atom information is still written to the textport with each individual pick.

The measurments for surfaces are Surface-Surface Distance, Surface-Surface Angle, and Surface-Surface Torsion. These work similarly to the above measurements for atoms. One can turn off the measurement function for the chosen button by reassigning it to Atom Information or Surface Information.

Scribing turns on the "scribing" function. This is a way to select parts of a surface by drawing the outline of an area onto the surface. Selecting this option turns the mode on and disables other functions of either button while the cursor lies over a molecular or accessible surface. When either the left or the middle button is held down while the cursor is over such a surface, and the mouse moved, the triangles under the cursor that make up that surface should "respond" by changing color to bright blue. Upon releasing the cursor, the "track" of the cursor should remain visible as a chain of bright blue triangles. Note that to move the surface while in scribing mode, one has to either use a dial box or position the mouse off of any surface before attempting to move the view by mouse.

When the user has outlined an area of interest with a blue border, the area can be "filled" by double­clicking anywhere inside the area. This will cause the initial triangle under the cursor to turn green, then the triangles adjacent to that triangle turn green, then those adjacent to those turn green, etc. Thus a green "wave" expands outward from the initial point. However, a blue triangle previously selected as part of the border will not turn green. Hence if the blue triangles form a complete circle about the selected area, the green expansion will halt inside the circle. If the border is not "water­tight", the green triangles will "spill" out and eventually cover the entire surface.

If the user clicks with either the left or middle mouse button while the wave is still expanding, it will immediately stop "growing". This can be used to highlight a region by double­clicking on the approximate center of the region, waiting for the wave to cover the desired area, then clicking again to make it stop.

The surface selection is now all the triangles colored green. This selection is referred to as the "Currently Scribed Surface" in all menus which prompt the user for a surface selection. For instance, "Calculate: Surface Area: Molecular Surface" path will give the user a menu with this option, which will result in the calculation of the surface area of the selection. The user can make this selection permanent by making it a formal subset. It will then be assigned a name which can be used to refer to it at any future time. This is described in detail under the Formal Subsets menu.

There are further options available if the user selects the Scribing option while scribing mode is already on: Initiate Fill, Undo Fill, Add Contiguous Border to Fill, Change All Border to Fill, Clear All Marks, and Turn Scribing Off. Undo Fill removes all green markings but leaves the blue ones. Add Contiguous Border to Fill changes to green only those blue areas which are next to green areas. Change All Border to Fill changes to green all blue areas. Clear All Marks removes all green and blue markings from the surface. Turn Scribing Off also removes all green and blue markings and also exits scribing mode.

One should exercise a little care when scribing for several reasons. First, if the cursor starts on the surface when a button is depressed but ends up off the surface before it is released (which should not really happen when scribing), one can get unexpected results (e.g. the mouse is "locked" into a rotational mode). Second, if the surface is at a coarse scale, e.g. a large molecule has been surfaced, then one should not scribe too fast because occasionally the program may get confused as to where it is on such a surface. Furthermore, if one is trying to trace a boundary on a complicated surface one may have to rotate the molecule to get over some surface features with the border intact.

Note that scribing does not work if the surface is rotated by dial while holding a mouse button down with the cursor on the surface. Also note that clicking on the same triangle twice causes the "greening" to begin, so be careful not to accidentally start this process by starting at the same site twice in a row. Finally, remember that, other than a well formed blue border, only a further mouse click will halt the expansion of the green triangles.

Command Line Mouse means the user can fix a certain written command to be automatically enacted when an atom is "picked". These commands can only refer to atoms at present. For instance, if the user enters "c=2" after selecting this item, then every atom picked after that will be colored 2. This procedure works by finding the atom number of the atom picked and then appending this to the entered command and sending it to the command interpreter. So if the user clicks on atom 55 then the command actually sent to the inperpreter is "c=2, an=55". Note this will not work with the projection command ("c=2 >r,an=55" will not color 2 the residue containing atom 55). One use of this function is to assign charges to a picked atom, or to remove certain atoms from view using "c=0" or "alt(r=0)". Choosing this entry again turns off the automatic command function.

Z­Trans. Alternatives affixes alternate functions to the Z­displacement mouse function, which is holding the middle mouse button down while moving the mouse up or down. There are three such functions: Z-Translate (Default), Z-Value of Pot. Plane, and Stereo Split/Twist. Z-Translate (Default) is the default and is used to reset this function. Z-Value of Pot. Plane alters the position of the projection plane, which is the plane colored coded by the potential from the current map. Stereo Split/Twist alters the separation of a stereo pair. This latter function is normally taken care of via the dial box, but is included since not all users will have one. Also in the latter case the stereo twist is fixed to the left­right motion of the mouse. This menu also returns the button to its usual functions.

4.5 - Read Menu

GRASP Surface File

DelPhi Potential Map

DelPhi Epsmap

GRASP Property File

PDB File

GRASP Script File

Radius/ Charge File (+Assign)

Curves File (.lis)

Pair Wise Interactions

Grasp .init File

Grasp Macro File

This menu deals with the input of data to Grasp, and the operations performed on it as a consequence. Formats for many of these files are described in Appendix A.

Upon choosing one of these options, the user will get up to three choices: a default file name, a menu list, or the option to enter a file name. When a file name is entered, Grasp checks first to see whether that file exists in the current directory. If it does not, then it then checks all the directories in the user's $PATH. If the file is not found, this will be reported in the texport and the read aborted.

To check the $PATH, type "echo $PATH". Grasp only checks $PATH at startup. The user can use this variable to point to any direcories which contain desired files. Some people have complained that Grasp has a hard time "seeing" certain files, claiming they don't exist when they do. If you have this problem, you may have $PATH set incorrectly. Even if Grasp is able to list (on a menu) files in your current directory, it will not be able to open them unless the first two characters of your $PATH are ".:" (dot-colon). If they are not, type "setenv PATH .:{$PATH}", or else type in all filenames in your current directory and prefix them with "./" (dot-slash). For example, a file in your current directory called lys.pdb would be typed as ./lys.pdb.

A Grasp Surface File contains the information necessary to reconstruct a surface. It is an unformatted, or binary, file and can not be read as an ascii file. It contains the surface scale and midpoint, plus a list of vertex coordinates, vertex normals, accessible surface points and a list of triangles, which are vertex numbers of each triangle. Also, any surface variable which has been calculated or assigned is also written to this file: surface potential, distance, curvature, and general properties 1 and 2. Upon reading this file, the surface is automatically displayed in the default surface mode. The surface display quantity is usually set for potentials. Since there are memory limits on the number of vertices and triangles current in the program at any one time, the program checks to see whether there is space for the new surface and the input is halted if there is not.

The program will write to the textport the number of vertices and triangles read, as well as which properties were also included in the file. Surface files may contain more than one distinct set of surface points, in analogy to a PDB file containing more than one molecule. Each surface read in is assigned a sequential integer, as if each had just been constructed anew - if the user has built two surfaces and two more are read in they are assigned surface numbers 3 and 4. These numbers can be used in surface subsetting. These numbers also appear in menu lists of constructed surfaces.

Grasp surface files should have the extension ".srf". They are one of the three primary data files for Grasp, by which is meant that the program can set the view center and scale, and can display the structure upon input.

A DelPhi Potential Map file, or phi map, can be produced by DelPhi or by Grasp. It is also an unformatted file and one of the three primary data files for Grasp and should have the extension ".phi". The program keeps a menu list of all such files in the current directory, which is accessible to the user when choosing a file. As well as containing the potential at every point of a 65 cubed lattice, phi maps contain a scalle (lattice spacing), and a mid­point (the absolute coordinates of grid point (33,33,33)). Extended phi maps may also contain a rotation matrix. A phi map is read into internal map 2, overwriting any resident map. This map is set to be the current map, and so is the one automatically used for such operations as contouring. If no scale and midpoint has been set for the Grasp unit box, it is set from the scale and midpoint of the phi map, so the map exactly fits the Grasp box.

A DelPhi Epsmap file (default extension ".eps") is the final unformatted file type Grasp will read and contains the information on the dielectric map used in a DelPhi calculation (eps is short for epsilon, the symbol for the dielectric value at each point in space). The dielectric map is defined at the grid mid­points, i.e. the points half way between every grid point and its six nearest neighbors. There are 3*65 cubed of these. Eps maps have a center and scale associated with them, though they can not be used as primary data files for Grasp. They also contain a list of which grid points were assigned as being in salt, which is referred to as the "Debye" map. When reading in an eps file, the user can choose to read in just the X or Y or Z component of the eps map, since these are each individually 65 cubed maps. The grid values are either set to one or zero, one if inside the low dielectric, zero if outside. The user can also opt to make a composite map of these three components, where the values of the eps map at each of the six midpoints associated with a grid point are added together and assigned to that grid point. Finally the user can select to read only the Debye map from the epsmap. Here the values assigned to each grid point are one if in salt and zero otherwise.

These maps can be used to check the actual surface used in a calculation, relative to the underlying atoms. They can also be useful in calculations. For instance, one can use the eps map to set all potentials inside the molecule to zero, or the Debye map can be used to calculate excess salt concentration from a DelPhi phi map.

A Grasp Property File is an ascii file which contains a list of values to be assigned to either atoms or vertices. The format of the file is especially simple, namely a number of comment lines, then a line with the letters "surface=property" if the values belong on a surface, or "atoms=property" if they belong to atoms, where property can be "potential", "distance", "curvature", "gproperty1", "gproperty2", "accessible", or "charge", followed by a list, one value per line, of the values to be assigned to vertices or atoms in sequential order - value 1 goes to atom 1 or vertex 1. When this file is read in, the maximum and minimum values for that property array are recalculated and the color coding altered if necessary if currently displayed. These files can be created by Grasp, or by the user via external programs.

A PDB File is the final primary data file for Grasp. The correct extension is ".pdb", and a list of all such files in the current directory is maintained in menu form. Grasp supports several variations on the format laid down by the Brookhaven Protein Data Bank, in that the field after the coordinates can be used to contain Grasp atomic property information. The original such variant was the "modified" PDB file output by DelPhi. This substituted the occupancy and B­factor information with radius and charge for that atom, since this is information was necessary and sufficient for a DelPhi calculation. Grasp supports this type of file and will recognize the format when the file is read and assign the values in these fields to the radius and charge of each atom. Grasp also supports files where this information is actually meant for general atom properties 1 and 2 (for instance if the values really do represent occupancy and B values), and where these properties are written in higher precision. To alert the program to the correct interpretation of these fields the user should insert the correct headers to these files, as described in section A.5. All of these file types are written by Grasp, in which case the correct header is automatically inserted.

Upon reading a PDB file, Grasp automatically constructs a bonding pattern for those atoms based upon proximity and atom type. This procedure is not always perfect but works well for high resolution structures. The exception to this is if the file contains alternate atoms, such as if some atoms are multiply represented in the file due to ambiguities in the original electron density map. The program will not at present take account of this in bond formation and inappropriate bonds will probably be made due to proximity considerations. Multiple structures can be entered within one file if they are separated by "TER" lines. Otherwise the same problem of inappropriate bonding patterns may occur. If Grasp detects what it thinks is such a pattern, like a carbon bonded to five atoms, it writes out that atom's information field, plus the number of bonds being made, to a file called bonding.dat. This can help the user check the validity of the assumed bonding by viewing this file after a structure has been read. Grasp will also write to the textport the number of atoms and molecules read.

Radii are assigned from the data file default.siz unless the file type has embedded radius information. Charges are not assigned automatically, unless they are also embedded in the file. If charges are assigned within the file the maximum and minimum charge values are (re)calculated for use in color coding of this variable. If no scale has been calculated, Grasp calculates the maximum X, Y or Z dimension and makes the Grasp box size 50% greater so that the molecule fills at most two thirds of the box. (Note that Grasp includes the atom radii in this assessment so that small molecules do not appear too large).

To do operations on DNA, it is necessary to read in a PDB file which contains DNA information. Mixed files with proteins and DNA are fine.

A Grasp Script File is an ascii file where that each line is read and sent to the command interpreter as if the lines were typed on the command line. As the file is read , each line is written to the textport at the same time it is processed. Lines which begin with the symbol "!" are treated as comment lines and written to the screen but not acted upon. An example of the use of such a file is to enact a default color scheme for atoms the user favors, or as an alternative method of setting radii or charges from DelPhi control files. The user has the option of writing all textport commands entered, via the Write menu, to an external file history.dat, which is often a good starting point in constructing a script file.

A Radius/Charge File deals with the input and processing of DelPhi style control files for these variables. Several such files are located in the Grasp data directory, and the user may place their own there too if they wish them to appear in the Radius/Charge File menu. These files should have the correct file extension: radisu size files are ".siz", charge files ".crg". Alternatively, the user can input the name of a file, which will be processed according to the file extension. Upon reading a charge file, the total assigned charge will be reported and the maximum and minimum values found, which may alter the display if charge is a current display variable. Upon reading a radius file, the program reports the number of distinct radii amongst all atoms. This value may not exceed 100.

A Curves File contains information on DNA parameters, and at present must be in the same format as that put out by the program "Curves", (which must have extension ".lis"). Future versions of Grasp will support other DNA structure program output.

A Pair Wise Interactions file is an ascii file which can come in two formats as described in Appendix A. These files contain information on which two residues each matrix interaction is between, and the strength of the interaction. If the program can not assign the given value to a pair and cannot find one or both of the described residues, that line of the file is written to a file called int.pair.not.assigned.dat. That there have been such failures is written to the textport after the file has been read in. There can be at most 10,000 interactions in the program at any one time. Note that reading in a new file will overwrite old values.

A Grasp .init File is an ascii file that contains one or more of the commands described in appendix B.

A Grasp Macro File is an ascii file that contains a series of definitions of commonly-used functions as described in section A.8.

4.6 - Write Menu

Grasp Surface File

Atom (PDB) File

Potential Map

GRASP Property File

History File

Surface Points as PDB

Macro to File

RGB/Snapshot File

Grasp can export most of its internal data structures to external files. The format of some of these files can be found in Appendix A. Files are written to the current directory (unless the file name specifies otherwise). If the file exists, the user will be prompted as to whether to overwrite the file or not. In some cases, the user may have the option to append to a file of the same name.

A Grasp Surface File contains information sufficient to display and manipulate a surface. Any property array which differs from zero in any of its entries is also written automatically to the same file. One can append surface files to other surface files when writing. When this is done the equivalent of a "TER" line is inserted so that upon subsequent input, different surfaces remain distinct, just as different molecules would from a properly constructed multiple­molecule PDB file.

When writing a surface file, the user is given the same choice of surface subsetting as in any other operation involving surfaces, so that one is not restricted to exporting whole surfaces. For instance, the user may wish to save only the part of a surface involved in making an interface with another surface. Note that the user can immediately read a surface just written, which provides a mechanism to duplicate a surface subset.

An Atom (PDB) File is a PDB file but with the four different possible formats mentioned in the Read menu section: no extra data, radius and charge data in the occupancy fields, another pair of properties in these fields, or a pair of properties to higher precision. In the latter two cases, the user is prompted as to which atom properties to export. As with surface files, the user may select any subset of atoms to be exported rather than all atoms, and the user has the choice of four different coordinate frames. As with surface files, the user can export a molecule or part of a molecule and then read it in as a way to duplicate those atoms.

For both surface and atom files, the user has four choices for the frame of reference used to calculate the coordinates written to file. These appear on a menu which has the entries: Absolute Centering, And Rotated, Box Centered, and And Rotated.

Absolute Centering, which is the default, is to write out the coordinates unchanged from the original PDB file. Box Centered writes out the coordinates after adjusting them so that the center of the molecule is (0,0,0). If a molecule has center (average) coordinates (4.203, 3.123, 5.011) and this is the first file Grasp reads in, it will set the scale of the Grasp box by it and also make it the center of the box. The box centered option subtracts 4.203 from each X coordinate, 3.123 from each Y coordinate, and 5.011 from each Z coordinate. The two And Rotated entries apply the current rotation/translation matrix to the original coordinates before writing. The first rotation entry applies just the rotation matrix, and the second applies the matrix and then centers the molecule at (0,0,0).

These options can be used to compare surfaces or molecules, or parts thereof, which have radically different centers and/or orientations. They can also be used to restore an orientation so that the exact same view can be examined again.

For Potential Map, Grasp will write out either internal map in DelPhi form. This file will contain the values at each grid point, along with the grid spacing and coordinates of the central point. Such files can not be concatenated. A default file name of fort.14 is provided because that is the default Fortran unit to which potential maps are written by DelPhi.

For a Grasp Property File, the user is given the choice of surface or atom property, and then the particular property to be written (if the user wishes to add comment lines to this file this must at present be done by hand - editing the file externally). One should note that the user does not get to choose which vertices and which atoms have their properties exported as all such are exported in Grasp property files. The reasoning behind this is that since the files do not contain any atom or vertex information other than the sequential position in the file, selecting a subset of, for instance, atoms, would not necessarily be sequential. Writing a property file for such a selection would create a file almost impossible to interpret, and which certainly could not be read back in. If a subset of properties is required, the user should export a property in a PDB file, or list the properties to to the textport with the "list" command and capture them using the cut and paste feature of the window manager.

The History File option will write all the commands processed by the program via the textport to the file history.dat. Note that only commands will be written, not selections passed to the program via the textport in the context of a menu request. The user can then edit this file and add comment lines by making the first character of that line a "!".

Cavity Surface Points as PDB will write a list of accessible surface points which belong to cavity surfaces (assuming these have been calculated) as if each such point was an atom, i.e. in PDB file format. The file is always named cav.pdb. One use of this file has been to add these pseudo­atoms to a pdb file and prevent this volume from being assumed high dielectric by DelPhi. (Note this is not assured of achieving this goal if the cavity could accommodate a sphere of diameter twice that of a water, but this rarely ever occurs).

Macro to File writes the definition of an already exisiting macro to a file. The user is asked for a filename and then to pick an entry from the Macros menu.

RGB/Snapshot File writes the currently displayed picture to an Iris "ipaste" rgb file.

4.7 - Formal Subsets Menu

Fix Dials to World

Fix Dials to a Subset

Make a Formal Subset

Remove a Formal Subset

Remove a Subset Rotation

Fix a Subset Rotation

Formal subsets may be a selection of atoms or surface vertices or both. If both, we refer to the subset as being of mixed type. Mixed type subsets come in two flavors - those where atoms were selected on the basis of proximity to a set of surface vertices, or those where surface vertices were selected on the basis of proximity to a set of atoms. Up to 100 formal subsets may be created. Upon creation, each subset is given a unique name by Grasp, or the user may provide a name. Formal subsets are formed to enable independent rotations and translations of a subset of a structure or to provide a unique label for a certain set of atoms and/or vertices.

The "world" has one transformation matrix associated with it and each formal subset has an additional unique such matrix. By "world" is meant the entire view including all atoms and vertices. The total transformation matrix for a subset is the product of these matrices. All subset rotations and translations of a formal subset change the subset's own private matrix, but have no effect on the world matrix. If newly defined subset is itself part of a formal subset which has been moved relative to the world, then the new subset will inherit the matrix of the previous subset.

Fix Dials to World and Fix Dials to a Subset allow the user to attach dials either to the world or to any formal subset. Once a formal subset is created and named, the dials are automatically assigned to that subset to the exclusion of all other atoms and/or vertices. If the user opts to connect the dials to another formal subset, the user is presented with a list of the subsets created so far. If active formal subset rotation mode (Control A) is in effect, the dials are still automatically attached to any new subset, however this is no longer significant, unless the user quits the mode immediately, since manipulations are only applied to what the cursor first lies upon. Note that choosing the world or a formal subset for the dials turns off the this mode.

Make a Formal Subset creates a formal subset. Grasp will prompt the user with the standard options for selecting a subset: command line selection, a particular molecule, a scribed surface, etc. These atoms or vertices then become the new formal subset. If the user want to make a mixed subset, the criteria used in associating surface with atoms or atoms with surface is distance. This can be distance in terms of atom center to vertex distance, atom center to vertex distance minus the Van der Waals radius of the atom, or "contact", which means that the surface and atom are related in that the atom was used in the creation of that bit of surface.

The names of formal subsets are derived from the constructed surface number of the surface, or the molecule number of the atoms, and any previously named subsets. For instance, if the user has selected atoms from the molecule 2, and it is the first such subset, then the name suggested would be "m2:a", "m" for molecule, "2" for molecule number, and "a" for the first such subset. If it was the second subset from molecule 2 and at least one atom in this selection is not contained within "m2:a", then the name suggested is "m2:b". If it is totally contained within "m2:a" then the naming becomes hierarchical and the name is "m2:a:a". If the atoms make up exactly all of one molecule then everything from the colon to the right is dropped, so that a subset containing all of molecule 2 is simply "m2". Note that molecules are not automatically formal subsets, they have to be defined as such. If the atoms are from more than one molecule then the "m" number is set to 0, so that the first such subset would be "m0:a", the second, if not included in m0:a, would be "m0:b", etc. The same rules apply to surfaces, where the surface number substitutes for molecule number. For mixed subsets, the rules for atoms with associated surface are applied as if there were only atoms, except the "m" becomes an "ms", making it "ms1:a" for the first set of atoms from molecule 1 with associated surface. For mixed subsets where the surface is the primary selection, and atoms are associated, the "ms" changes to "sm", so that the set of some vertices from surface 2 with all atoms within five Angstroms would be "sm2:a".

This somewhat complicated naming system was designed to enable the user to keep track of complicated subsetting operations. If the user is not going to require such sophistication, choosing an arbitrary name may be preferable. Note also that if the subset created contains elements of a previously created formal subset, those elements are reassigned to the new subset.

One can also Remove a Formal Subset. When this is done, the atoms and/or vertices formally associated with that subset are returned to the world - its own private transformation matrix is wiped clean, and the formal subset name is removed from the name list. This may cause the subset to "jump" if this subset has been moved relative to the world. If a surface subset has been made which does not consist of a complete surface, for instance the user scribed a patch of a complete subset and made this a subset, then deleting a subset does not return the internal representation completely to its original state. This is because forming such a subset will cause the duplication of some vertices, specifically those at the border of the subset where vertices are originally shared with the rest of the surface. This may cause problems if the user later attempts to scribe across this region into another, or in any other Grasp function where vertex connectivity is important.

Remove a Subset Rotation allows selective "undo" operations on rotations and translations applied to subsets. One of the intentions of formal subsetting was to allow the user to rearrange the positions of two molecules, or parts of molecules - for instance to explore new configurations of an enzyme and an inhibitor. The menu options for removing and fixing subset rotations are designed to help this process.

After selecting a subset, the user has three choices: That Subset Whole Rotation, Since World Last Moved, and Since Another Subset Moved. That Subset Whole Rotation will remove all the "differential" manipulations on the chosen subset and resets the subset's own matrix to the identity matrix. Hence the subset is moved to the position it would have had if it had never been selected as a formal subset. Since World Last Moved removes partial rotation and translation operations applied to the subset since the world was last moved, and Since Another Subset Moved removes all operations such since some other formal subset was moved. These last two options reset the subset's matrix to previously saved values, e.g. that matrix when the world was last moved.

Fix a Subset Rotation means that the subset's own transformation matrix, the matrix which represents the differential manipulations relative to the rotations and translations applied to the world view, is applied to the subset's internal coordinates. Its values are then reset to the identity matrix. Remember that moving an object via mouse or dials does not affect an object's internal coordinates - just the view. This option allows the user to "fix" the apparent rotation, make the change of view apparent in the objects coordinate's, and reset the view. This application of a transformation is described in Write menu under PDB File. Note that if the user chooses to fix the world, then all coordinates are recalculated, and world transformation is set to its initial value, the identity matrix.

If the user is in stereo or split screen mode, subset manipulations can become very complicated. Grasp allows the user to manipulate the left and right views independently. This facility carries over to formal subsets, so that one can manipulate the right­hand view of a formal subset independently of the left­hand view. To do this, merely set the dials to the right­hand view ("Display: Alter Stereo/Split: Dials to Right") and attach the dials to the desired formal subset with Fix Dials to a Subset. To switch to the same subset on the left, do the same thing for the left­hand view. Note that being in active subset rotation mode does not help here, since the side which is manipulated is NOT decided by which side the cursor initially resides upon. Furthermore, not all features are fully implemented, for instance the "undo" features of Remove Subset Rotations only apply to the left­hand subset. This limitation also applies to Fix Subset Rotations. It is unlikely this will present many problems to most users. This will be resolved better in later versions.

4.8 - Programs Menu


This menu is intended to allow the user to access auxiliary programs via Grasp. At present the only such program is superimpose, which uses the Kabsch algorithm to calculate the r.m.s between sets of atoms.

Superimpose requires two set of atoms as its input. The user can supply these in the usual manner, formal subset name, command line selection, complete molecule, etc. These two sets MUST have the same number of atoms. Once given, the algorithm calculates the rotation and translation which will minimize the root mean square difference between these two sets, the differences being calculated between each atom in each list sequentially, atom one of list one with atom one of list two, etc. This r.ms. value is reported via the textport.

The user is then given the option of actually superimposing the two sets of atoms or larger subsets of atoms based upon the same rotation and translation. For instance, suppose one is interested in the similarity of two helices within a protein. First, select an equal number of residues within each helix. Since the residues of each helix may not be exactly the same, the residues can not be automatically compared or superimposed. So select just the backbone alpha carbons from each helix. An r.m.s is then calculated and reported for these two subsets.

The user is then presented with a submenu to apply the rotation matrix. No quits the superimpose program. Apply Forward to 1st Selection rotates and translates the atoms of the first set onto the second via the optimal matrix calculated. Apply Backward to 2nd Selection will do the same but with the second set onto the first. In the example above, this is not a very useful operation in either direction - one wants to compare entire helices, not just the alpha carbons. Apply Forward to New Selection and Apply Backward to New Selection both apply the calculated matrix to a new selection. Choosing the whole of the first helix and apply forward to new selection, or the second helix and apply backward to new selection, will superimpose the complete helices.

4.9 - Set Parameters Menu

Rotation Rate

Translation Rate

Probe Radii Values

Electrostatic Parameters

Depth Cueing

Material Transparency

Worm Parameters

System Miscellaneous

DNA H-Bond Parameters

This set of menus gives the user access to many of the internal parameters of Grasp. The menu contains entries for parameters often altered and submenus for parameters which can be grouped together.

Rotation Rate and Translation Rate alter the rate at which the view is rotated and translated by the dials or the mouse. The user enters new values via the textport. To leave the value unchanged, hit return without typing anything. Rotation rates are in angles per redraw, while translation rates are in Angstroms per redraw. The default values in Grasp are set high to allow for rapid initial manipulations, after which the values can be set smaller for fine control.

Probe Radii Values brings up a submenu which accesses the radii of the different probes used in Grasp, namely those used in calculating surface area of atoms, making a molecular surface, and in a Poisson­Boltzmann calculation (the latter can also be set by Electrostatic Parameters). The same probe is used for both molecular surface and accessible surfaces.

Electrostatic Parameters allows the user to alter the values which are used in the Poisson­Boltzmann calculation when the user selects "Calculation: New Potential Map". Those parameters are the inner and outer relative dielectrics, the water probe radius used to estimate water inaccessibility, the probe size used to account for ionic radius, which is how close an ion may come to the surface of a molecule, and the ionic or salt concentration. The default values for these parameters are:

interior dielectric 2.0

exterior dielectric 80.0

water probe radius 1.4

ionic probe radius 2.0

salt concentration 0.0

Depth Cueing allows the user to control the apparent depth of a view by having parts of a view further away appear darker than parts closer. In theory one does not have to interpolate colors to black, but that is the automatic choice in Grasp. Colors are recalculated based upon a front distance f and a back distance k (f > k). If a vertex has Grasp Z­coordinate of z and a color of (r,g,b) its depth-cued color (r',g',b') is found from:

r'=r*F, g'=g*F, b'=b*F where F=(Z­k)/(f­k), (f­k)=max(0.1, f­k), and (F=1.0 if F> 1.0) and (F=0.0 if F<0.0).

In Grasp, f is taken from the front of the object and k is defined relative to it as a multiple of the distance from the front to the back of the object. So if an object has a front to back distance of 0.8 box units and a depth factor of 0.5, the back of the object has its colors half-interpolated to black. If the depth factor is 0.0 then the back distance is infinity and depth cueing is turned off. A depth factor greater than 1 will result in some of the object being colored completely black (the element is not uncolored, it's black). The front and back of the object in question, whether a surface such as a contour or molecule surface, or for bonds or atoms, is calculated dynamically and changes as the molecule is moved. Note that the last condition specified above, (f­k)=max(0.1, f­k), is included so that depth cueing does not become too excessive for small molecules. In general the larger the molecule the more depth cueing is needed, the smaller the molecule the less is needed. Another way of stating this is that the degree of depth cueing should be proportional to the complexity of the object.

The user gets the option of entering a new depth cueing value which is the inverse of k, or turning the depth cueing off for atoms, bonds, molecular surfaces, contours, or all structures. Depth shading is highly recommended for surfaces drawn in a mesh representation.

Material Transparency allows the user to set the material transparency for a molecular surface, contour, worm, interpolation plane, or all of these. The transparency options are 3/4 solid, 1/2 solid, 1/4 solid, or opaque (default).

Worm Parameters changes the parameters used in constructing a backbone worm. These options are Worm Thickness, Spline Resolution, Cross Section Resolution, and Allowed Spline Gap. Spline Resolution alters the number of subsegments used for each B­spline segment. Each segment is constructed from each set of four consecutive atoms in the selection list, and the default number of subsegments per segment is four. Note that this change only takes effect when a worm is built and does not alter the segment density in chains already constructed.

Cross Section Resolution alters the number of sides of the cross-section of the polygonal tube used to construct each segment. The default number of sides is ten.

Worm Thickness alters the radius of the cross-section of the worm. Note that changes made to thickness and segment resolution are reflected immediately in any worm already built.

Allowed Spline Gap is the distance used to determine spline breakage. If two sequential atoms in a worm selection are more than a certain distance apart, the spline is terminated and a new one begun. Hence, if constructing a backbone worm when the protein has more than one chain, there will be one spline per chain. If this distance is set sufficiently large, this will force the construction of a single spline through disjoint chains. It can also be useful in judging patterns of distances of a certain subclass of atoms - spline breakage contains local distance information.

System Miscellaneous brings up a submenu with the entries: CPK Level, Surface Area Probe Density, Bond Cylinder Diameter, and Maximum Surface Resolution. CPK Level allows the user to select the resolution used to draw the solid spheres used to represent atoms. The different levels represent hierarchical decimations of an octahedron.

Surface Area Probe Density controls the number of points on the probe sphere used to calculate surface area in Grasp implementation of Shrake and Rupley's algorithm, where points on a sphere of radius=(Van der Waals of atom)+(radius of water) are tested as to whether they lie within the similarly expanded radius of any other atom. The fraction of such points which do not is the fractional accessibility of that atom from which the accessible area is derived by multiplying by the area of that atom's expanded sphere. The point densities are based upon decimation of an icosahedron. On choosing a level, the number of points in the test sphere is written to the textport.

Bond Cylinder Diameter is the thickness of the cylindrical bond representation in Angstroms. The user can enter any value between 0.0 and 1.0. Default is 0.2 Angstroms.

Maximum Surface Resolution corresponds to the minimum lattice spacing allowed for the lattice used in creating surfaces. The smaller this number, the finer the grid spacing, the higher the resolution allowed. This number is not allowed below 1/3 Angstrom. Making this number larger forces a coarser grid to be used, which results in fewer triangles and surfaces which are quicker to spatially manipulate in real time.

DNA H-Bond Parameters allows the user to set the DNA H-bond parameters.

4.10 - Miscellaneous Menu

Toggle Cross Hairs On And Off

Toggle Color Bar On And Off

Release/Reconnect All Dials

Fullscreen/Normal Toggle

Set And Save Colors

Smooth Property

Z-Slice Tool

Invert Normals

Change Current Map

Reset World Rotations

This menu has functions which do not quite fit anywhere else. Some of these entries can be accessed via control keys. Most entries are self­explanatory and so we shall describe only those which are not.

Smooth Property smooths the interpolation of surfaces.

Z-Slice Tool invokes a special tool which allows the user to control the clipping planes for the view. These clipping planes apply to all structures, including the display box. The tool appears as an independent window, i.e. it can be placed anywhere, resized at will and quit like any other Iris window. However the implementation is not perfect and at this time it is recommended that the tool is NOT resized and that the user be VERY careful not to quit the entire program when exiting a Z­Slice window (there are two options here, "close" and "quit", always choose "close").

The tool appears as having four colored squares surrounded by a white border. This border will change to red if the cursor is placed over the tool. Each square controls one aspect of the slice plane, namely either the width, the mid point, the front plane position or the back plane position. Each square has its function inscribed along with the current value. These values are in Angstroms. By clicking and releasing either the left or the middle mouse button in a square the value associated with that box is altered. Clicking to the left of the center of a box decreases the value, to the right increases it. The closer to the center a click the smaller the increment. The right most mouse button brings up the windows control menu. When the tool is first activated, the clipping planes are +/­1.0, i.e the front and back of the Grasp box. It is recommended that the user have this box displayed (Control O) while using the Z­Slice tool since this clarifies the clipping planes' position.

Invert Normals inverts the surface normals of a surface selection, which is a way to make a surface appear "inverted" or "inside out".

Change Current Map swaps the pointer to the current internal map. This can also be accessed via Control C.

Reset World Rotations erases all "world" rotations and translations and returns the view to when the first structure was read into Grasp.

4.11 - Help Menu

The help menu has two entries, Command Line Syntax and Control Keys. Command Line Syntax displays a list of Grasp commands, each with a short description. Control Keys displays a list of control Keys and their functions. Selecting an entry in the Control Keys menu causes that function to execute.

4.12 - Macros Menu

This menu displays the defined names of all macros which have been read in with "Read: Grasp Macro File". Selecting an entry on this menu cuases that macro to execute.

4.13 - Quit GRASP

"It's Miller time".

5 - Command Line

Most menu commands can be entered from the keyboard instead. When entering commands from the keyboard, the cursor should initially be over the graphics window. The user then just begins to type the command. The cursor will then jump to the textport where the characters will appear. Upon completion of the command, hit 'return' and the command will be executed and the cursor will return to the graphics screen.

(The textport may originally not be in the foreground and has to jump from the background. This usually takes a finite length of time to complete and if the user types particularly fast the second character entered may get lost in the transition and will not appear in the typed command. If the textport is in the foreground this usually does not occur. Also if the user tries to backspace over the first letter of the command they will find they can not! Simply reenter the command in this case.)

Commands that can be entered via the keyboard are color assignments for all structures, precise rotations and translations, radii and charges of atoms, listing of atom properties to the screen, information for atoms and surfaces, and subsetting codes for commands and for menu­driven functions. Note that Grasp is case insensitive for character variables, such as atom names, but not for subsetting codes.

To simplify the description, the following notations will be observed for variables:

n,m any integer

x,y any real number

a,b,c,d any character

5.1 - Subsetting Codes

Commands usually include subsetting codes. One can subset by many properties of atoms, surfaces, and other objects.

5.1.1 - Negation, Ranges, and Concatenation in Subsetting Codes

Subsetting codes are negated by putting a minus sign right after the equals sign:

r=-lys selects all residues which are NOT lysine.

If the variable to be negated is a number, put parentheses around it to avoid confusion:

q=-(1.0) selects all atoms which do NOT have a charge of 1.0.

q=-1.0 selects all atoms with a charge of -1.0.

Ranges are selected by separating range values with commas and putting them in parentheses:

p=(1,2) selects all atoms whose potential is in the range 1.0 kt to 2.0 kt.

Subsetting codes are concatenated by separating them with commas:

r=lys, q=-(0.0), X=<0.0 selects all atoms which are in lysines AND have non­zero charge AND are in the left hand side of the screen.

5.1.2 - Atom Subsetting Codes

Atom Name: a=abcd

a=oe1 selects all atoms with the name "oe1" or "OE1" or Oe1" or "oE1".

There are up to 4 characters read for each atom name.

Question marks act as wild cards:

a=oe? selects all atoms with names "OE1" and "OE2".

"_" indicates an intentional blank:

a=o_1 all atoms with names "o 1" or "O 1".

There are a couple of short cut names for backbone atoms and for side chain atoms:

a=ba selects atoms C, CA, N, O, HA, and HN.

a=sch equivalent to a=-ba.

Atom Number: an=n, an=(n,m), an=>n, an=<m

an=(1,500) selects the first five hundred atoms.

Atom number refers to the internal numbering scheme - atom 256 is the 256th atom to be read in, NOT atom 256 in the pdb file.

Residue Name: r=abc

r=lys selects all atoms in all lysines.

There are 3 characters read for each residue name.

There are several short cut names for residues based upon their hydrophilicity:

r=crg selects all residues which are normally assigned formal charges - lysine, arginine, glutamate, aspartate.

r=pol selects all the residues formally polar - serine, threonine, tyrosine, histidine, cystine, asparagine, glutamine, tryptophan.

r=hyd selects all hydrophobic residues - alanine, valine, leucine, isoleucine, methionine, phenylalanine, proline.

Residue Number: rn=n, rn=(n,m), rn=>n, rn=<m

rn=(1,25) selects all residues which have residue numbers between 1 and 25.

Unlike atom numbers, residue numbers are the same as in the pdb file. A consequence of this is that while every atom has a unique number, different residues (e.g. on different chains) could have the same number. If a residue number is not assigned in the pdb file it is assigned the number 1 for internal purposes.

Residue Projection: >r

Projection allows selection of a residue based upon whether an atom of that residue has been selected. Suppose one wants to select all residues which have atoms within five Angstroms of a certain cofactor with residue name CYT. First, one calculates a distance map for all atoms to this cofactor. One would then select all atoms which are within five Angstroms of the cofactor and "project" this onto the parent residues:

d=<5.0,r=-cyt >r selects all residues with atoms within 5 Angstroms of residue CYT.

Chain Name: ch=a

ch=-B selects all atoms NOT in chain B.

If there is no chain specifier in the pdb file, it is assigned the letter "A".

Charge: q=x, q=(x,y), q=>x, q=<y

q=>0.0 selects all atoms which have been assigned a positive charge.

Radius: R=x, R=(x,y), R=>x, R=<y

R=-(0.0) selects all atoms which have been assigned a non-zero radius.

Potential: p=x, p=(x,y), p=>x, p=<y

p=>10.0 selects all atoms at whose center the calculated potential is less than 10.

Potentials are in kt, (1 kt = 0.6 kcals).

Original Coordinates: x=x, y=(x,y), z=>x, z=<y

x=(50.0,60.0),y=>45.0 selects all atoms whose original x coordinate is from 50 to 60 Angstroms and whose original y coordinate is => 45.

These are the coordinates as they appear in the pdb file, in Angstroms. These are unchanged by any user applied rotations or translations.

Screen Coordinates: X=x, Y=(x,y), Z=>x, Z=<y

X=>0.5 selects all atoms whose current screen x coordinate is => 0.5.

These coordinates are relative to the screen and in box coordinates. They alter as the molecule is moved. The coordinates here are relative to the unit box, i.e. which runs from plus to minus one, so the example given above will pick all atoms which are in the right most quarter of the screen.

General Property 1: p1=x, p1=(x,y), p1=>x, p1=<y

General Property 2: p2=x, p2=(x,y), p2=>x, p2=<y

p1=(5.5,6.5) selects all atoms whose general property 1 is from 5.5 to 6.5.

Molecule Number: m=n, m=(n,m), m=>n, m=<m

m=1 selects the first molecule that was read in.

Molecule number refers to the order in which molecules are read in, i.e. the first structure read in is assigned the number 1, the second 2 and so on. If more than one molecule is read from a single file, the molecule numbers are sequentially assigned.

Accessible Area: S=x, S=(x,y), S=>x, S=<y

S=0.0 selects all buried atoms.

One should first do an accessible area calculation to be able to select on it.

Distance: d=x, d=(x,y), d=>x, d=<y

d=<3.0 selects all atoms 3 Angstroms or less from some set of points.

One must have calculated a distance map to select on this property.

Formal Subset Name: sub=abcd . . .

sub=m1:a selects the formal subset "m1:a".

If you choose your own formal subset names, rather than accept those produced for you by Grasp, try not to start the name with a "-", since this will be interpreted as a NOT symbol. Subset numbers can also be substituted for formal subset names (i.e. if m1:a was the first subset created, then sub=1 will have the same affect as sub=m1:a).

Discrete Atom Color: cd=n

cd=7 selects all atoms assigned to color 7.

5.1.3 - Surface Subsetting Codes

Many of the subsetting codes for surface properties are the same as those for atoms. In context there should be no danger of non­uniqueness. When using the vertex coloring command "vc=n, p=n" it is clear that the potentials being specified by the "p=n" code refer to vertices and not atoms. Similarly any requests for subsetting via the menus will entail a similar understanding. The following codes are identical for both atoms and vertices (they both refer to the same property but for atoms in one case and vertices in the other):

p= potential

x=,y=,z= orginal coordinates

X=,Y=,Z= screen coordinates

p1=,p2= general properties 1and 2

d= distance

sub= formal subset name

Surface Number: s=n, s=(n,m), s=>n, s=<m

s=(1,3) selects the first three surfaces read in or constructed.

Surface number refers to the order in which surfaces are read in or constructed, i.e. the first surface read in is assigned the number 1, the second 2 and so on. If more than one surface is read from a single file, the surface numbers are sequentially assigned.

Curvature: C=x, C=(x,y), C=>x, C=<y

C=>0.0 selects all parts of the calculated surface which are concave.

One must first calculate the curvature to select on this property.

Vertex Number: vn=n, vn=(n,m), vn=>n, vn=<m

vn=15677 selects vertex number 15677.

Vertex number is assigned serially upon being imported or constructed. Vertex number might not seem a useful variable, but for example if one calculates the maximum potential on a surface, or portion of a surface, the vertex number of that point is also returned. Hence a "vn" code along with a "vc" (vertex color) command can be used to locate this point.

Discrete Vertex Color: vcd=n

vcd=2 selects all parts of the surface currently assigned color 2.

5.1.4 - Bond Subsetting Codes

One selects bonds on the basis of the atoms they originate from, or the atom they are ending on. Note that although bonds conceptually go between pairs of atoms, in Grasp they go halfway - each bond object goes from the center of an atom halfway to the other atom of the bond pair. Each half-bond is then uniquely associated with an atom. So when a bond is colored, actually only half the bond is colored.

Bond colors can be automatically mapped from underlying atoms via the appropriate menu command. There is also an internal color scheme for bonds which can be selected from the menu.

Bond Projection: >p

a=sch >p selects all bonds ending at side chain atoms.

Usually one selects bonds by selecting a subset of atoms. The bonds which originate from those atoms are then selected. Bond projection is used to "project" to a subset of atoms the bond is ending on:

Discrete Bond Color: bcd=n

r=pro,bcd=3 selects all bonds in any proline currently assigned color 3.

5.1.5 - Backbone Box Subsetting Codes

One selects backbone boxes by selecting a subset of atoms. The backbone boxes are constructed out of four atoms: the alpha carbon of the first residue, the carboxyl oxygen of the first residue, the amine nitrogen of the next residue and the alpha carbon of the next residue. If any of these atoms are colored, the backbone box that uses that atom is assigned that color.

The only unique property backbone boxes possess is the color they have been assigned:

Discrete Box Color: kcd=n

rn=(8,33), kcd=3 selects all backbone boxes which are part of residues 8 to 33 which are currently assigned the color 3.

5.1.6 - Pair-Wise Interaction (Matrix) Strand Subsetting Codes

Matrix strands are residue-based but they can be subsetted using any atom codes. If then any atom of a residue is selected then that residue is selected. Both ends of the strand are checked for and ORed to determine selection. There is no direct way to select on the basis of both ends of the strand, for example only strands which go between lysines and glutamate (this can be done indirectly using the icd code). This is an exception to the usual Grasp philosophy of commands always being AND based and reflects the two­site nature of matrix strands.

All other matrix strand operations, such as scaling by distance, deciding upon the display mode, and the maximum interaction strengths used in width encoding, are under menu control.

Strand Strength: ip=x, ip=(x,y), ip=>x, ip=<y

ip=>0.0 selects all strands with positive interactions.

Strand Strength Rank: ip=n

ip=2 selects all strands which are the most positive or second most positive for that residue.

ip=-1 selects only those strands which are the most negative for that residue.

Discrete Strand Color: icd=n

icd=2 selects all strands currently colored 2.

This code can be used to do the "double" selection mentioned above - select all strands running between lysines and glutamates. The following set of commands will color only these 2:

ic=0 uncolors all strands

ic=3,r=lys colors 3 all strands originating or ending on lysines.

ic=2,icd=3,r=glu colors 2 all strands originating or ending on glutamates which are already colored 3.

ic=0,icd=3 uncolors all strands which are still colored 3.

5.2 - Coloring

The commands for coloring are:

c=n change atom to color n

vc=n change vertex to color n

bc=n change bond to color n

kc=n change backbone box to color n

ic=n change matrix strand to color n

wc=n change backbone worm segment to color n

where n can be between 0 and 99 and indexes the Grasp color set. Color 0 is always black and coloring anything 0 will cause it to disappear.

The default color for atoms and bonds is 1 (white). The default color for surfaces is 91. The color 1 is not used because it is usually altered to be less than pure white, which although is okay visually for atoms, looks awful on surfaces, whereas color 91 is always unaltered white. The default color scheme for backbone boxes is white at the alpha carbons, red at the oxygen, blue at the hydrogen to indicate the electrostatic character of the peptide plane.

There are several exceptions to the color argument being an integer n:

c=u "undo" the last atom color command.

vc=u "undo" the last vertex color command.

These commands will change the color of each atom/vertex changed in the last command to what it was before. Whenever atom or vertex colors are changed, a backup copy is made of the original array and this is retrieved if an "undo" is entered.

c=r "restore" the last set of atoms.

vc=r "restore" the last set of vertices.

These commands act upon the atoms/vertices which have most recently been set to color 0 (hidden), and restores their color to what it was before they were hidden. While "undo" will only undo the last coloring command, "restore" will keep restoring until all colors are changed back from 0. Although these commands are often used without selecting a subset, they can also be subsetted to "undo" only part of a color command or "restore" only part of a hidden subset.

vc=a transfer atom colors to associated vertices.

This command transfers atom colors to the associated vertices. By "associated" is meant the atom­to­vertex mapping provided by the intermediate accessible surface. This can be subsetted only by atom codes. This is the only exception to a general "like­with­like" rule for atoms and surfaces - that only atom properties can subset atoms and only surface properties surfaces.

kc=d reset backbone boxes to their original default colors.

This command restores the coloring of the selected backbone boxes to the default - corners white, red, or blue depending on the atom.

ic=c work out the connectedness of all visible strands.

This command causes Grasp to work out the connectedness of all visible strands. That is to say if two strands will be assigned to the same group if they are connected to the same residue, or if it is possible to go from one to the other via other visible strands. Grasp will then report the number of such patches and color each differently. (Only nine different colors are used so if there are more than nine patches the colors repeat). Note that there is no restriction applicable to this command - one can not restrict this command to certain residues. Instead one would first color all strands which are greater than a certain strength, then issue the 'ic=c' command. This will then show how far "webs" of that interaction threshold spread.

Examples of coloring commands:

c=7 colors all atoms 7.

bc=2,r=pro,bcd=3 colors 2 all bonds in any proline which are currently colored 3.

bc=4,a=n??? colors 4 all half-bonds originating from any nitrogen atom.

kc=4,rn=(8,33),kcd=3 colors 4 all backbone boxes which are part of residues 8 to 33 which are currently colored 3.

ic=0 makes all matrix strands disappear.

vc=a, ch=b maps the atom colors associated with chain b onto the associated surface.

ic=0, icd=2 undisplays all matrix strands currently colored 2.

ic=2, ip=>0.0, a=oe1 colors 2 all strands which have positive strength and originate (or end) on a glutamate or glutamine since these are the only residues with atoms labelled 'oe1'.

ic=2, ip=1, r=lys colors 2 all strands which are the most positive strand coming out of either residue it connects, and for which either of these residues is a lysine.

wc=2,rn=(8,33) colors 2 all backbone worm segments belonging to residues 8 to 33.

wc=0,q=-(0)>r undisplays all worm segments which belong to residues which have at least one atom charged.

5.3 - Precise Rotation and Translation

Sometimes it is necessary to move objects by exact distances or angles of rotation. These commands can be supplemented only by formal subset names. For example, "xr=90, sub=m1:a" will rotate the atoms and/or vertices of formal subset "m1:a".

xt=x translate in the x direction by x Angstroms

yt=x translate in the y direction by x Angstroms

zt=x translate in the z direction by x Angstroms

xr=x rotate about the x axis by x degrees

yr=x rotate about the y axis by x degrees

zr=x rotate about the z axis by x degrees

Xt=x translate in the x direction by x box units

Yt=x translate in the y direction by x box units

Zt=x translate in the z direction by x box units

5.4 - Listing Atom Properties

The list command prints atom properties to the textport for each atom selected. Without subsetting, this command will list properties for all atoms. If there are more than 4 atoms selected, Grasp will automatically expand the textport to list up to 20 atoms at a time. If there are more than 20 atoms in the requested list, hitting the space bar will give the next 20 atoms, while hitting return lists the next 1 atom. The output can be terminated by hitting any key other than the space bar or return. Upon completion of the listing, hitting return reduces the textport to its original size and position.

list, r=lys,a=nz lists information for all terminal zeta nitrogen of lysines.

The properties listed are controlled by the information parameter list. The default properties for atoms are atom name, residue name, residue number and chain name, while for surface vertices it is potential. Every possible property for surfaces and atoms is assigned a single letter code:

Atom Code Surface Code

default atom information a original coordinates x

original coordinates x surface potential p

screen coordinates X curvature C

distance d screen coordinates X

radius r distance d

charge q surface number s

potential p formal subset name b

molecule number m cavity number V

general property 1 1 general property 1 1

general property 2 2 general property 2 2

formal subset name b

One can alter the list by adding properties, removing properties, or resetting the whole list:

si=pC reset surface list to ONLY potential and curvature.

ai=qr reset the atom list to charges and radii, but do not remove default atom information if it is on the list already.

ai=+b add formal subset name to the atom list.

ai=-a remove default atom information from atom list.

si=+xd add absolute coordinates and distance to the surface list.

Note that the order of the parameters entered does not change the order in which the parameters are written to the textport. That order is determined by the position in the above table of properties from top to bottom, which corresponds to left to right output.

5.5 - Altering Radii and Charges

The commands to change radii and charges are:

alt(r=x) change radius to x

alt(q=x) change charge to x

Commands to modify existing radii and charges are:

alt(r=r+x) modify radius to x plus existing radius

alt(r=r*x) modify radius to x times existing radius

alt(q=q+x) modify charge to x plus existing charge

alt(q=q*x) modify charge to x times existing charge

alt(q=1.0), a=nz, r=lys will assign a plus one charge to all zeta nitrogens of lysine.

Charge is a display property for atoms, so if the charge distribution is altered, it will cause a recalculation of the maximum and minimum values used for color scaling.

One can use these commands in an external file as an alternative method of assigning charges and radii to the DelPhi control files. Simply edit a file to contain the list of commands to assign charges or radii, or both, and then read in the file as a Grasp script file (section A.7).

Atoms of zero radii are not displayed or used in calculations of curvature, accessible area, or low dielectric volume.

5.6 - History and ! Commands

History commands print selections of the history list, which is a list of all commands entered via the text window.

history n (or his n) causes the last n commands to be written to the textport.

The default value of n is 10, so that if n is not entered, the last 10 commands are written to the textport. History commands are NOT added to the history list. Only commands which are sent directly to the command interpreter appear on the history list. Selections typed during menu-driven actions do not appear.

! commands allow the user to reexecute previous commands stored in the history list.

!! (or !) causes the last command to be reexecuted.

!n causes the nth command to be reexecuted.

n causes the nth previous command to be reexecuted.

!­1 causes the previous command to be reexecuted.

All reexecuted commands are added to the history list. An example of the use of such commands would be to incrementally rotate a molecule about an axis:

xr=2.0 rotates about the x axis by 2 degrees.

! rotates about the x axis by another 2 degrees.

! does it again.

Adding the characters ":p" to a ! command causes that command to be printed but not reexecuted or added to the history list. For instance,

!­1:p prints the previous command but does not reexecute or add it to the history list.

Editing history commands is not yet implemented.

6 - Control Keys

A control key in Grasp means any alphabetical character depressed while the "Ctrl" key is depressed. To activate control keys, the cursor must be on the graphics window. They are included because people (for instance me) find them useful to have as an alternative to menu­driven functions. Most control key functions can also be found in menus.

Control A - Toggle Active Subset Rotations On/Off

If Grasp is manipulating more than one formal subset object independently, only one such object can be "attached" to the dials or mouse­dials at one time. This can be set via the menus. However, if one wants to switch back and forth between different subsets continuously this can be frustratingly slow. Switching to active subset rotation mode means that when the user is using the mouse to enact translation or rotations the object moved is that upon which the cursor lies when the mouse button is first depressed. Essentially the object is "picked", then moved. If the cursor lies over no object at the beginning of the motion, then the mouse­dials are attached to the world and everything is moved together. If dials are used instead of the mouse then the object moved is either the one last moved by the mouse or the one initially "attached" to the dials when the active mode was invoked. Turning active subset rotation mode off (pressing control A again) leaves the dials permanently attached to the last moved object. Note that this mode is turned off when the world or a formal subset is chosen via the Formal Subsets menu.

Control B - Toggle Color Scale Widgets On/Off

Color scales should automatically pop up when a continuous coloring scheme is being displayed. If they do not, or if the user wants to remove them from view, they can be turned on and off with control B. The color scale has two components, the title space, from which a menu may be accessed by the right mouse button, and the rest, which controls the color coding.

The color coding part should consist mostly of a colored section and a small white section, with the symbol ">­<" on it, at the right­hand end. The colors should be the colors in use for that property and structure, e.g. red, white and blue for electrostatics, and should vary continuously from right to left. If the coloring mode is three color continuous, the colors will vary from the first to second to third colors, if two color continuous, they will vary from the first to third color. Printed on this colored strip should be five equally spaced numbers which increase from left to right. The left, middle and right numbers are those used in the color coding. If the cursor is placed over any of these three numbers and the left mouse button pressed and held down, that number will decrease and continue to decrease while the button is held down. If the middle button is used, the number will increase. When the button is released, it causes the display to update colors based upon this new number.

If the scale is in three color mode and the value being altered is the lowest value, the rate of increase or decrease depends upon the difference between the lower value and the middle value - it increases by a constant fraction (10%) of that difference. Similarly the upper value changes based on the difference between the upper and middle values. The middle value changes based on the difference between it and the value towards which it is being altered. In two color mode only the left and right values change, and they do so based on their difference.

The ">­<" part of this widget allows compression and expansion of the color scale range. Depressing the left mouse button and holding it on this symbol is equivalent to increasing the left number while simultaneously decreasing the right number. The middle button will expand the range by decreasing the lowest number and increasing the highest number. This is often useful in viewing electrostatics since the range of potentials is usually much wider than is useful for distinguishing positive and negative parts of a surface, so compressing the range will improve the picture.

The color scale menu allows the user to enter new values for the control (i.e. minimum, middle, maximum) numbers, also RGB triplets for the colors used. It also allows the user to reset the control numbers to their original values. There is also support for changing the draw mode and property displayed. Two color continuous is invoked by setting the middle value to the lowest value, and since values can only be altered by fractions of a difference, one must use this menu to so set the control values, and to reset to three color continuous. The color bar menu also allows the user to alter the colors used for display, the draw mode for the surface or atoms, and the property being displayed. These last two options produce menus identical to those which appear in regular menu use for these features.

One color bar should appear for each different quantity displayed - if the user is displaying atom potentials and surface potentials, two bars will appear. If the user is using the split screen mode and different properties in the left view and right view, one scale will appear for each.

Control C - Change Current Map

The current map is the 65 cubed set of grid values which are used to build contours, interpolate potentials at a slice plane, interpolate potentials at a surface etc. There are two such maps in Grasp, the first of which is the default space for all internally generated maps, and the second for all maps read into Grasp. Hitting Control C produces a menu so the user can choose which is "current". The user should beware of hitting Control C when the cursor is NOT over the graphics window, since if it is over the textport the program will terminate.

Control F - Toggle Full Screen View On/Off

This expands the Grasp graphics window to fill up the entire screen and returns it to the window view.

Control L - Free/Fix Light Source

The direction from which the light source produces lighting effects on rendered surfaces can be altered after hitting Control L. Moving the mouse (without depressing any button) will then move the light source in that direction. (The source is set at infinity and so only the direction of the light source is altered) Hitting Control L a second time fixes the light source's new direction. Note that this will affect the lighting of ALL surfaces, including those pseudo­lit and those which make up the surface of "CPK" atoms. The user should experiment since different lighting angles often bring out different features of surfaces.

Control M - Toggle Textport Depth

This will pop the textport to the front if it is lower down, and push it to the back if it is in front.

Control O - Toggle Grasp Box On/On/Off

The Grasp box represents a box of +/­1, in screen coordinates, in each direction. The first Control O produces a box which is depth shaded, i.e. the box sides get darker the further from the viewer. The edges are also outlined in black. One advantage of this view is that because it has simulated depth it can fool the the eye into expecting any other object in view to have depth. Hence it "trains" the eye to see the objects in the box as three dimensional. Pressing Control O again removes the depth shading of the cube, leaving all sides white. This is provided in case the user is capturing pictures to send to a postscript printer (it makes a difference). Hitting Control O once more removes the box.

Control P - Bring Up The Color Palette

The Color Palette is the tool with which users may alter color from within the program. What should appear when Control P is hit are nine colored squares, each one with a quadrangle of white, red, green and blue attached to the lower, left, upper and right sides respectively. The colors shown inside the squares are the first 9 of the 99 indexed colors of Grasp. The index number of each color is written in the center of each square.

When the tool is first invoked, the colors displayed are:



















































The next nine colors are accessed by hitting the space bar, and the next nine after that by hitting it once more, and so on. After colors 91 to 99, it returns to colors 1 to 9. Colors 10 to 89 are left intentionally blank for the user to create their own. Colors 91 to 99 repeat colors 1 to 9.

To alter a color, the user positions the cursor over one of the side quadrilaterals of that color, then either the left or middle button is depressed. The middle button increases that component, the left button decreases that component. If the cursor is on the white component it increases/decreases all components. Colors are updated within the squares as the button is held down, as are the red, blue or green component values. The user will probably find that uniformly decreasing the red/green/blue components of colors 1 through 9 from their above given values will give richer display colors.

Altered colors have their RGB triplets automatically stored in the file "defcol.dat" in the current directory. This file is automatically read in the next time Grasp is started within the same directory. When colors 91 to 99 are changed, their new values are not saved, so these are always the same as the default colors 1 to 9.

When the user is finished, hitting any key other than the space bar, will remove the palette from the screen. Any structure colored by a color which has been altered will automatically update its color.

Control Q - Quit

Control R - Toggle between Single and Double Buffered Mode

Animation is achieved on an Iris by drawing a view into a secondary or "back" buffer, then switching it into the primary or "front" buffer only when the drawing is complete. In this way none of the actual drawing process is seen. Iris machines come with a limited amount of hardware for these two buffers. Power series machines have two 24­bit buffers allowing "full" color mode for animation. Most lower level machines come with one 24­bit buffer which is split into two 12­bit buffers for animation. Having 12 bits for three colors means that there are only 4 bits per color, rather than 8 in full mode. Hence the same variety of colors are not available for animation. If the user has such a system and wants to get a 24­bit picture they should switch to single buffer mode. Then all 24­bits are used for coloring, but one "sees" the drawing as the view is constructed. The best use of this is then to enhance a view the user is not going to change, e.g. if one is going to take a photograph from the screen. And occasionally it may be instructive to see how a view is drawn.

Control S - Stereo/Split Screen Mode Menu

This menu is described in section 4.1.

Control U - Unhook Dials

If the dials are used extensively, the user may find one suddenly ceases to work. It has come to the end of its range. If this happens, Control U will turn off the dials so that the useless dial can be rewound without moving the view. Pressing Control U again turns the dials back on.

Control V - Repair Surface

Occasionally upon construction of a molecular surface there may occur "defects" in the structure - holes in the rendered surface. These can usually be fixed by hitting Control V once or twice. This is best done before another structure is constructed or imported.

Control W - Swap buffers

Swap the current front buffer for the back buffer (see Control R).

Control X - Toggle Cross­Hairs On/Off

Sometimes the cross­hairs are not useful. This turns them off (and back on again).

Control Y - Toggle Projection Plane

The projection plane is like having a molecular surface stretched across the Z=0 plane from X=(­1,1) and Y=(­1,1). The potential at any point in this plane is calculated from whichever potential map is current (although, of course, the values in the maps do not have to be potentials) by the usual trilinear interpolation. Because the plane is linked to the Grasp box, it does not move when the view is rotated, hence the map position of each point on the plane will change, since maps are rotated with the view. Hence the potentials are recalculated upon every time the view is moved. The color coding is calculated as if each point in the plane indeed belonged to a molecular surface, i.e. using the same colors and control values. Hence they may be altered the same way, via the color scale menu. One advantage of the projection plane is that by moving the molecule back and forth in the Z direction, one can see patterns of potentials within the molecule. It is also possible to move the projection plane rather than the molecule, by altering the Z value from the default of 0. This is achieved via the "Z­Trans. Alternatives" option on the "Mouse Functions" menu.

Control Z - Unix Shell

This launches a new Unix shell so that the user can issue any unix command, edit files, change current directory, etc. Of course the user can always do these things from another window, but sometimes it's just more convenient to hit Control Z. To exit the shell, type "quit" or "exit" or "logout". Grasp is "frozen" until the shell is successfully exited. This temporary shell does not have access to any of user's login aliases, nor will it understand the "~" symbol, or other short­cuts which are set up for the user's usual shell.

7 - Worked Examples

In all the examples described below it is assumed a PDB file or other primary data file has been read in and is currently displayed.

7.1 - Color a molecular surface by electrostatic potential

First assign charges. When PDB files are read in, the charge on each atom is set to 0, unless charge information and the correct header were included in the file (section A.5). The default charge set for proteins is be found in the file full.crg. Read this in and assign charges with "Read: Radius/Charge File (+Assign): full.crg". The total charge assigned is reported in the textport. Note that histidines are uncharged. Atoms can be charged by hand with the "alt(q=x)" command. If a "partial" charge set is employed, where atoms other than ionizables are charged, then there is often a problem of incomplete charging due to inconsistent naming conventions of atoms. If a residue has a non­integer charge after a charge assignment from a file, this is reported to the textport and the file charging_data is written to the current directory with information on such residues. Remember that you can write pdb­like files with the charge and radii information for any subset of atoms (section A.5).

Having charged the protein, consider if the default values for an electrostatic calculation are what you want. These are changed by "Set Parameters: Electrostatic Parameters" and the default values are listed in section 4.9. Next proceed to launch the Poisson­Boltzmann solver with "Calculate: New Potential Map". This should take a few seconds to complete, upon which you have filled map 1 with potential values.

Next construct a molecular surface with "Build: Molecular Surface". At the completion of this procedure, a white molecular surface should appear. The surface will appear in the default display mode which is a fully lit surface unless otherwise set in the file .init_Grasp. Change the surface display mode with "Display: Alter: Molecular Surface: (Color Surface by): (Surface Display Mode)" (the ( ) indicate that no choice was made from these menus and the existing or default coloring is being used). Note that the atoms are still being displayed, although this may not be apparent if the surface is opaque. Since the display of the atoms will slow the draw speed and make the surface appear to have "lines" in it, you might want to get rid of them with "Display: Hide: Atoms".

"Calculate: Pot. via Map at Surfaces/Atoms" will calculate the potential values at all atoms and all surface vertices from map 1. This will produce faint colors (red and blue) on the white surface. Also the color scale should appear in the top part of the box. If it does not, hit Control B. Use this to alter the color coding. The most useful action will probably be to place the cursor over the ">­<" part of the color bar and hold down the left button until the right and left numbers are less than 10 in absolute values. The color saturations will then be stronger.

7.2 - Display surface potential and curvature side by side

Having calculated the surface potential in the previous example, now calculate the curvature. Do this with "Calculate: Surface Curvature (+Display)". A menu will ask you two questions: which surface and what atoms. Since there is only one surface, that part is easy. As to what atoms should be chosen, this depends upon the set of atoms used in the construction of the surface. As a good general rule, use the same set of atoms in calculating the curvature.

Upon completion of this calculation (note that this is NOT the fastest calculation in Grasp), the surface quantity will switch to curvature. This should be obvious since the default color scheme for curvature is green/grey instead of red/blue. Also the title of the color scale will change from "Potential" to "Curvature". Adjust color coding to the desired intensity.

Next change to split screen mode with "Display: Stereo/Split On: Dials to Both". There should now be two surfaces color coded by curvature. Move these apart a suitable distance if necessary (use the bottom right dial OR use "Mouse Function: Z­Trans. Alternatives: Stereo Split/Twist"). Then do "Display: Alter: Molecular Surface: Potential: (Surface Display Mode): Left". This last will change the display of the left-hand surface to be by potential. Note that two color scales are now visible, one for potential, linked to the left view, and one for curvature, linked to the right view.

7.3 - Surface two interacting parts of a molecule and select the interface

Create two formal subsets from two sections of the molecule with "Formal Subsets: Make a Formal Subset: Atoms: Enter Specifications" and select any subset of atoms you wish according to the rules in section 5.1. Accept the suggested subset names "m1" and "m2". Surface each subset individually with "Build: Molecular Surface: A Formal Subset: m1", then "Build: Molecular Surface: A Formal Subset: m2: Yes" (the second surfacing will ask you whether to add to or overwrite the first). Having built two surfaces, we want to display, or color, only those parts of the two surfaces which are proximal to each other. To do this, select "Calculate: Distance Array: A Surface to a Surface: A Constructed Surface: Surface 1: A Constructed Surface: Surface 2". Note that this calculation is the slowest in Grasp so be patient! This will calculate the minimum distances from each vertex of the first surface constructed to any vertex on the second surface. This calculation only results in values being calculated for the first surface and so the reverse calculation also has to be done with "Calculate: Distance Array: A Surface to a Surface: A Constructed Surface: Surface 2: A Constructed Surface: Surface 1" to find minimum distance values for the second surface.

Now that distance is a property for both surfaces, you can remove all but the interfacial regions by giving the color command "vc=0,d=>x", where x is whatever you want. For instance, if x=5.0, then only parts of the surfaces which are within 5 Angstroms of the other surface will remain visible. You can make distance the display property of the remaining portions of the surfaces with "Display: Alter: Molecular Surface: Distance: (Surface Display Mode)". To bring back parts of the surface that are hidden, type "vc=1".

7.4 - Calculate the occluded accessible surface area between these parts

Following from the previous example, select "Calculate: Area of a Surface/Molecule: Molecule: Accessible Area: A Formal Subset: m1". This will result in various text appearing in the textport. At the end of this will be a number, the total accessible area for that set of atoms. Write this down. Now "Calculate: Area of a Surface/Molecule: Molecule: Accessible Area: A Formal Subset: m2". This will give the accessible area of the second set. Write this down too.

Now we want to calculate the accessible area of subsets m1 and m2 in the context of each other. The difference between this area and the sum of the areas for m1 and m2 separately will give the occluded accessible area between m1 and m2. It may not be simple to select the set "m1 and m2". One way to do this is with the series of commands: "c=1", "c=2,sub=m1", "c=2,sub=m2". Now all atoms are colored 1, except in subsets m1 and m2 where all atoms are colored 2. Now the code "cd=-1" will select all atoms that are NOT colored 1.

Now "Calculate: Area of a Surface/Molecule: Molecule: Accessible Area: Enter String" and type "cd=-1". The number in the textport at the end of this is the area. Add together the numbers you obtained from the first two calculations and subtract the third number from the sum. This is the occluded acessible area between m1 and m2.

7.5 - Display atomic B­values on the surface

Edit a PDB file containing B­value information such that the first two lines read:



This tells Grasp to read columns 55-60 and 61-67 and store them in general properties 1 and 2. Upon reading this modified file, the B­values will be stored in general property 2. To see the atoms displayed such that they are color coded by B­value, select "Display: Alter: Atoms: Property #2: (Atom Drawing Mode)". A color scale should appear which you can use to improve the color coding, alter colors etc.

Now build a molecular surface for the whole molecule or some interesting subset of it. After you have done this, select "Calculate: Simple Property Math: Map Atom Value to Surface: All Surfaces: All Atoms: General Property #2: Potentials". This will project the B values into the surface potential array of Grasp, which can then be colored and displayed independently of the underlying atoms. Note that if instead of "All Atoms", you select a subset which does not include all the atoms used in creating the surface, some of the surface vertices will not be assigned a value. Values are only transferred from atoms in "contact" with the surface.

7.6 - Calculate and display effective dielectric from a single charge site

The "effective" dielectric at a site is defined, for our purposes, as the ratio of the potential at the site due to a charge at a second site when there are two dielectric values, as opposed to when there is just one - it is the ratio of potential for a 2/80 Poisson­Boltzmann calculation to that for Coulombs Law for a dielectric of 2.

First we have to charge a single site. Instead of using an "alt" command we will illustrate the use of the "mouse command line" function. Select "Mouse Functions: Command Line Mouse", then enter "alt(q=1.0)". This connects the function of assigning the charge of +1 to the act of picking an atom. You can now select an atom on the molecule and charge it instantly. To remove this function, repeat "Mouse Functions: Command Line Mouse".

Next calculate a new potential map with "Calculate: New Potential Map". Then store this map in map 2 with "Simple Property Math: Potential Maps: Map1 <-> Map2". Next set the external dielectric to the internal dielectric by selecting "Set Parameters: Electrostatic Parameters: Outer Dielectric" and entering "2.0". Then calculate the potential map again. Now the uniform dielectric results are in map 1 and the two dielectric results are in map 2. We want to divide map 1 by map 2, which we do with "Calculate: Simple Property Math: Potential Maps: (Map) op (Map) = (Map): Divide Maps: Map1: Map2: Map1"., which divides map1 by map2 and puts the results in map 1. Map 1 now contains "effective" dielectric values for all its grid points. Selecting "Calculate: Pot. via Map at Surfaces/Atoms" then interpolates these values at every atom and vertex. These can then be displayed as usual.

Note that we are using a uniform dielectric grid calculation to estimate the Coulombic potential at every atom/vertex. Since there is only one charge this could also be done by calculating a distance map and then using the "simple math" utility a couple times, along with the fact that a charge in dielectric of two produces a potential of 280.5 kt one Angstrom away. Considering the inaccuracy involved in the two dielectric calculations, this is hardly worth it. The values calculated should definitely be treated with caution for this reason, especially close to the charge where grid effects will be largest. However this is a calculation worth doing at least once to get a feel of the effect of two dielectrics on the potential distribution, i.e. of the distance at which solvent screening takes effect.

7.7 - Calculate surface area by hydro-phobic and -philic residue

First calculate the total surface area of all atoms with "Calculate: Area of a Surface/Molecule: Molecule: Accessible Area: All Atoms". This fills the internal atom array for accessible area. Next choose "Calculate: Simple Property Math: Atom Properties: Enter String", enter "r=hyd", then "Sum of Values: Accessible Area". The will result in the printing of the total area of all hydrophobic residues. Repeat the above and type "r=pol" for hydrophilic residues. Note that we do not want to choose "Average Value" here because the average is over atoms, not residues. To find how many hydrophobic residues there are, try coloring the alpha carbons of each residue with enter "c=1,a=ca,r=hyd". The number of atoms colored will be the number of such residues. Similarly for hydrophobic residues. The residues defined as "hydrophobic" or "hydrophilic" are described in section 5.1.2 under Residue Name.

7.8 - Find and display all residues within 3 Angstroms of an active site

The cleanest way of selecting an "active site" is to scribe it. This method is described in section 4.4 under Scribing. Once you have done this, the surface of the active site is green. The next step is to create a distance map for all the atoms with "Calculate: Distance Array: Atoms to Surface: VdW Surface to Surface: All atoms: Currently Scribed Surface". This will calculate the nearest surface vertex of the bright green subset to each atom, and subtract from this distance the atom's radius.

Once this calculation is complete, we want all residues within three Angstroms. Since we only have atom information, we want to use the residue projection code described in section 5.1.2 under Residue Projection, so type "c=2,d=<3.0>r" so that all atoms of all residues fulfilling this criteria are colored 2 (red). Now hide the surface to see the red atoms around the active site.

Note the command c=0,d=>3.0>r" will "uncolor" all atoms of all residues which have at least one atom at a distance greater than three Angstroms, which is NOT the complement of the previous command.

7.9 - Find the common volume between two superimposed molecules

This example requires that you have loaded two fairly similar molecules. Section 4.8 describes how to successfully superimpose two molecules. To find the common volume after this, select "Build: Consensus Volume". This creates a grid map where a value of 0 means outside of both molecules, 1 means inside of either molecule but not both, and 2 means inside of both. This map is stored in map 2. We want to contour this map, so we need to ensure map 2 is the current map. Do this with "Miscellaneous: Change Current Map: Inputted Delphi Map (Map2)". Next choose "Build: Contour: 3-D", and enter "2.0", then "4". This should produce a blue contour which represents the boundary of the "consensus" volume. The final step is to calculate the volume of this contour with "Calculate: Volume of a Surface/Molecule: Contour Surface: Value= 2.0" which returns the volume of this contour in cubic Angstroms.

7.10 - Form a six-helix bundle from a single helix

First select a helix of sufficient length from a larger molecule. Write these atoms to an external file, exit Grasp and restart it with this new file. We want to reproduce this helix. The simplest way is to read in the file five more times. This results in six identical helices, all exactly on top of each other, but with different molecule numbers. Align these helices so that the long axis is along the Z axis, into the screen.

To manipulate the helices independently of each other, we need to make a formal subset of each one. A quick way to so this is with "Formal Subsets: Make a Formal Subset: Atoms: Make All Molecules Subsets". The program will then prompt you with its suggested names, which will be "m1", "m2"..etc. The helices are now formal subsets. The dials are attached to helix 6, since this was the last subset created. Move this helix to the right until it is clear of the other helices. Then attach the dials to the world with menus "Formal Subsets:Fix Dials to World" and rotate the view about the Z axis by 60 degrees. This might be more easily accomplished by the rotation command "zr=60".

Then attach the dials to helix 5 with "Formal Subsets: Fix Dials to a Subset: m5", move it to the right until clear of the other helices, then fix the dials to the world, rotate 60 degrees about the Z axis, fix the dials to helix 4 etc. The end result should be that all six helices are separated and roughly at the corners of a hexagon.

Next change to active subset rotation mode by hitting Control A. Now helix moved will be the one the cursor lies upon. Adjust each helix to a "comfortable" position relative to the other helices in this mode. Remember that the world view can be moved by starting with the cursor off of all molecules.

When you are happy with the arrangement, the relative positions of the helices should be "fixed" so that the arrangement is not lost. Remember that the internal coordinates of the molecules have not been changed yet, and it is these that most functions within the program use, such as surfacing and the Poisson­Boltzmann solver. To fix the coordinates, select "Formal Subsets: Fix a Subset Rotation: m1", then "Formal Subsets: Fix a Subset Rotation: m2", etc., until all are fixed. Note that all the helices are still formal subsets. You might want to remove each subset after fixing it, with "Formal Subsets: Remove a Formal Subset: m1", and so on. This only removes the name of the subsets and rejoins them to the "world" object. This prevents a user from spoiling a careful arrangement by accidentally moving a subset.

At this point you can perform any Grasp operation as if the assembly has just been read into the program, for instance create a molecular surface, do an electrostatics calculation, write the new coordinates to a PDB file, etc.

8 - Future Developments

8.1 - General Improvements

The following is a digression into what I hope is installed within the program in the next six months, even though software completion is unpredictable.

The drawing of surfaces in Grasp is done in a per triangle basis, i.e. one triangle at a time. While this approach simplifies the drawing of partial surfaces, it is not as efficient as the per mesh approach. This means that collections of connected triangles are drawn together, taking advantage of the SGI mesh subroutine calls. S. Sridharan has produced general meshing algorithms which increase drawing rates by a factor of 2.7 compared to a theoretical maximum of 3.0. This will speed the drawing of any complex surface, i.e. molecular, contour etc.

Rex Bharadwaj has been responsible for the DNA objects within Grasp. At the moment it relies upon external programs to calculate DNA parameters. It would be easier to generate some parameters internally, i.e. the ones most often used like base pair twist and tilt. There will also be support for other DNA parameter scales than those provided by CURVES. An interface will be constructed which allows the user to display the results of dynamical simulations in object form. Reference forms of DNA will be included for comparison purposes. The ability to deal with multiple structures, "pick" objects and return values plus access to Grasp tools such as the color scale will also be included.

Residues will be given a more independent status, i.e. similar to that atoms and surface vertices have at present, e.g. having properties distinct, though usually derivable, from the underlying atoms. Instances of such properties might be residue dipole, residue area, residue volume, distance to other residues, phi and psi angles etc. This will facilitate mapping of residue properties onto residue objects, i.e. color coding by property, and onto surfaces. It will also allow the user to write and read residue property files.

Local averaging and differencing of properties will be introduced for surfaces and atoms and maps. This will allow for smoothing of properties and also the calculation of new ones, e.g. field strength for maps. Cross­surface distances, i.e. as opposed to Cartesian distances, will be added as a property.

A "math" interpreter will be written which will allow for more complex mathematics on surface and atomic properties. This will allow for conditional operations on sets of data which are hard to perform within the program at present. A simple graphing utility may be added.

Hydrogen bond representation and calculation will be added. As an object these will have intrinsic properties, e.g. angular and distance differences from "ideal" structures, as well as inheriting the properties of the atoms forming the bond. As with matrix representations, the user will be able visualize hydrogen bonding networks, i.e. hydrogen bonds which share atoms, or residues, or chains.

S. Sridharan has written much more efficient algorithms for the calculation of surface areas and also the curvature of those area elements. These will be interfaced to Grasp. He also has algorithms for the prediction of ion binding sites based upon electrostatics. These will be improved with desolvation and hydrophobicity penalties and added to the program.

Generic objects will be introduced to represent sets of atoms, e.g. cylinders, spheres. These may also be included as low dielectric volumes in Poisson­Boltzmann calculations. Properties of underlying atoms and residues may then be inherited by these objects and displayed thereon. Map properties may also be interpolated at the surface of such objects. These objects can be thought of as tertiary structure representations in analogy with those for secondary structures.

Reduced representations of surfaces will be introduced. Atoms and groups of atoms can be represented as spheres, worm lines, backbone boxes etc. In keeping with the duality within Grasp of vertices and atoms similar simplifications will be made for groups of vertices.

Grasp originally had animation facility i.e. the ability to display successive frames of a simulation based upon Discover (molecular dynamics from Biosym Technologies) output. It was removed until a better interface for this feature may be developed. This may include animation of reduced representations in analogy with that for DNA structures.

The secondary structure type of each residue will be assessed, either by previously published criteria or by internally adjustable constraints (e.g. hydrogen bonding patterns, phi and psi angles etc).

Material surface properties will be adjustable. The number of properties accessible to .init_Grasp will be greatly increased. A global rescale will be included. Previous commands will be accessible via a "history" command as in Unix. Text labelling will be added.

Bugs will be fixed.

8.2 - Intelligent Delphi

Intelligent DelPhi (Id) will automate many of the tasks performed by DelPhi, taking unto itself the task of organizing multiple runs and collating relevant data. Parameters such as dielectrics, salt concentration etc. would be set in a "DelPhi Panel", independent of the particular use. Those uses are outlined below.

The question of accuracy, where appropriate, would be settled via an option for "high", "medium" or "low" precision. These range would refer to accuracy ranges of roughly 0.1 kt, 0.25 kt and 1.0 kt for the particular quantities required.

Obviously more accurate calculations would require more time. Id would indicate the expected duration of the calculation before running, provide regular (visual) updates on the progress of the calculation, as well as options to abort the calculation.

Delphi is not always necessary or sufficient for all functions. For example there is a requirement for algorithms for Van der Waals energies, surface area, and molecular volume. Hence Id is also a superset of DelPhi functionality.

All functions which produce single number answers, such as total solvation energy, will produce break downs of these numbers to include, for instance, the effect of salt, if present in the calculation, and any other subsets which go into a composite calculation.

The user will have the option of saving results at various levels of detail to files. Also log files and files used temporarily by ID will be available for the user to track if they wish, or in the unlikely case of a crash, to find out what went wrong.


1) Find potentials in a cubic box.

This would be similar to what is done now in Grasp except that the user could specify the size, orientation and position of the box.

This box would be visualized in Grasp and its parameters altered by a widget. Alternatively these parameters could be entered explicitly.

The size could be via the molecules size (i.e. percent fill), or absolute size (e.g. 100 Angstroms). The molecules size can be defined either by the maximum dimension in the X, Y or Z direction, by the maximum length of the principal axis of the molecule, or to the largest separation between any two atoms.

Orientation can be determined relative to the principle axes of the molecule, or determined by the direction of maximum separation of any two atoms, or by any two points the user cares to define.

The central position could be by atom set or surface set, or entered explicitly. If the DelPhi box boundaries are too near the molecule focussing runs would be automatically queued.

All other DelPhi parameters, such as salt concentration, dielectrics, etc. would be entered explicitly, with defaults for all values.

2) Find the potentials at a given set of points given a certain charge set.

These points would usually, though not exclusively, be at atom coordinates. This charge set could be the charges of all atoms or a subset of atoms.

Results can be incorporated into Grasp, or exported to external files. Potentials can be broken down into contributions from permanent charges, polarization and from mobile ions, or given as total electrostatic potentials.

3) Find the interaction between two sets of charges.

To some extent this is a subset of 2), i.e. given a set of charges find the potential at a second set of sites. The work of extracting the interaction energy from this type of calculation is done for the user. One of the uses of this would be to find the effect of one ionizable residue upon another. Another would be the electrostatic interaction between two molecules.

4) Total solvation energy.

By this is meant the total electrostatic energy involved in the creation of a dielectric discontinuity between molecule and solvent.

Because of the nature of the algorithms used it is often much less computationally expensive to find the difference in solvation energy for two or more molecules, or conformations of a single molecule, than to calculate the individual values for each. Hence the user will have the option to specify a set of structures and take advantage of the improved speed of these difference calculations.

5) Construct an Interaction Matrix.

Use function 3) repeatedly to find the interaction between all members of a set of residues. Then use 4) to find the energy of charging a residue on its own. The former produce the off­diagonal and latter the diagonal elements of an interaction matrix which can be used in ancillary algorithms to estimate effective PK's of residues.

6) Total transfer free energy.

The total electrostatic energy of transferring between solvents, plus a contribution from volume effects, plus a contribution due to hydrophobicity.

7) Total Binding Energy.

Total electrostatic energy between two sets of atoms plus a contribution from Van der Waals energy and hydrophobicity.

8) Helix / Membrane Affinity.

Membrane affinity based upon an approximate desolvation penalty, plus hydrophobicity, per residue in the context of the primary sequence as an alpha helix.

9) Surface Area, Curved Surface Area, Van der Waals energy, Coulombic Energy.

This can be used individually on any molecule or subset of a molecule. Output can be contracted to give totals, or assigned as properties within Grasp.

Programs and Grasp Features

1) Facilities to make changing and applying charge sets easier.

This is often the most time consuming part of DelPhi, as currently implemented, because of non­standard naming conventions for atoms.

2) De novo charge sets.

For new structures, e.g. drug molecules, it will be important to have a charge set of some kind. Doree Sikoff has Scheraga's program implemented which will act as a first step in this direction.

3) Proton placement.

Marilyn Gunner's program to assign positions of protons based upon geometric constraints.

4) PK analysis.

Programs from Gunner and Yang to estimate effective pK's of ionizable residues.

5) Solids.

Facility to include geometric shapes in the dielectric calculation.

8.3 - Secondary Structure Display

Why another secondary structure program? After all there are some excellent representation programs available, e.g. RIBBONS by Mike Carson. Specifically because I think there is a lack of emphasis in such programs on secondary structure representation representing anything except structure. Grasps philosophy of representation is that as well as representing structure, objects must also be able to represent properties of the underlying atoms or vertices, and may also have intrinsic properties. Hence elements of a secondary structure representation should also be able to have their shape and/or color altered by underlying properties such as hydrophobicity, potential, and B­values as well as intrinsic ones such as twist, curvature, and strain. The following is the envisioned implementation of Grasp's secondary structure package.

Each residue will have a position and a direction associated with it. The position, or point, might be the atom center of the alpha carbon, or the average backbone position, or the middle of the side chain. It might also be the the closest point on the axis of a "perfect" helix defined relative to this residue. The direction might be the along the C­C beta bond, i.e. pointing towards the side chain, or might be the direction to the next residue, or to the nearest non­neighboring residues. Both the point and direction definitions can be user implemented with a considerable degree of flexibility.

Given a set of points, i.e. those belonging to a set of residues along a chain, a smooth line can always be drawn. Popular "splines" are the B­spline because of its inherent smoothness, and "Cardinal" splines which pass through control points. Each residue can then be assigned a line "segment". (In essence this is a property of the difference in positions of the points belonging to sequential residues before and after each residue). This line segment has dimension one as opposed to that of the point for that residue which has dimension zero.

The residue line can be expanded into a sheet, or ribbon, of two dimensions given a direction. This direction could be that assigned to the residue, or could be derived from properties of the line itself, i.e. the second derivative of the line at each point.

Given a direction associated with each line segment one can also define a twist, i.e. the variation of this vector as a function of distance along the line. The variation of this quantity is an interesting property in its own right. It should be constant for perfect sheets and helices, and random for true random coil.

The residue line can also be expanded to three dimensions into a tube. This is the most common representation seen today. The tube can be circular in cross­section, i.e. having no directional character, or be ellipsoidal in which case there is directionality in the long axis. This latter can again be correlated with the intrinsic residue direction or with that from the line segment. Circular tubes have one quantity i.e. thickness, ellipsoidal tubes have two, i.e. the lengths of the minor and major axis.

As described above we have four logical values we can assign to each segment, namely it can be a line, a ribbon, a circular tube or an ellipsoidal tube. For instance we can assign lines to random coil segments, circular tubes to helices and ellipsoidal tubes to beta sheets. Furthermore the ellipsoidal tube and the ribbon can display directional properties of a residue and, along with the circular tube can represent at least one property by via its size. Each element has as intrinsic properties of direction, a rate of change of direction, a twist and a rate of change of twist (higher derivatives may or may not be meaningful) either from the form of the line or from the intrinsic residue direction. Each element also inherits all the properties of the underlying residue. Any such scalar variable can be represented as the width of the line structure or as a color as with surface or atom colors. Any vector property can be set to be the intrinsic residue direction.

Hence this approach will give each residue an object i.e. the line segment, which comes in several flavors and which can represent both vector and scalar properties as well as having intrinsic properties.

One interesting example of the intrinsic properties is the variation from ideality of a helix or sheet along its length. This is often a difficult property to visualize but, as occurred with curvature, the quantification and color coding of this property will probably be highly instructive.

8.4 - Docking with Realistic Energies

The docking of molecules, either by hand or by some automatic procedure, is of particular interest because of the importance of shape (i.e. surface) complementarity. As has been described earlier Grasp allows the user to "invert" a surface. When this is used for interacting surfaces it allow the user to match "like­with­like" rather than "knobs­with­holes". However, the physical docking of surfaces or molecules is hard to do manually. One project for Grasp is to improve the visual aids to a user attempting to do just that, and also to provide numerical feedback, for instance distance calculations and energy analysis in as close to real time as possible.

Graphically many of the tools are already present, for instance the independent manipulations of objects and the split screen capability. However these can be greatly enhanced with some additional features. One example would be "proximity strands". Given a certain set of points on one subset one might want to know the nearest set of points on the subset being moved relative to it. This can then be graphically illustrated by drawing strands between the pairs of points. These could then be dynamically updated as the user moves the subsets relative to each other. The split screen functionality can be used to great affect here. If the subsets are arranged on the right in an orientation such that the interface regions of both are clearly visible, while the dials are attached to the left view, the strand positions could be updated based upon the apparent left view arrangement, but displayed on the right, were they may be more clearly seen. The clarity of view can be further enhanced beyond just a different relative arrangement on the right­hand view. If one is looking at a "deep" binding site, e.g. a small molecule into a deep cleft, then on the right­hand view the user could split the cleft into two parts along its axis and "peel" each part back, i.e. exposing the "interior". This then further avoids any loss of visual information due to local topology, i.e. the small molecule obscuring the cleft.

Another approach which would help in docking molecules is to allow rotations and translations to be preformed in a "local" coordinate frame, i.e. one linked to the molecular shape, rather than the XYZ system of the screen. For instance in docking of an elongated molecule into a deep pocket of another one might want to assign one axis pointing into the axis. Such local coordinate frames can be confusing in general and so a good compromise would be to allow the user to switch back and forth between a "local" and a "global" frame.

Speeding up distance calculations to the point where they can be done in real time would also have a useful graphical impact. At the moment such algorithms in Grasp are quite slow but by using certain grid techniques it ought to be possible to speed this approach greatly. Similar comments would then apply as for proximity strands to the uses of a split screen approach.

Energy analysis can be split into four classes of energies, namely Coulombic, Van der Waals, Hydrophobic and Desolvation. The first two terms are traditional in the sense that they are included in packages available elsewhere.

The hydrophobic contributions can be assessed, at least according to theories developed in this lab, as being proportional to the excluded accessible area, where elements of this area are weighted by their accessibility to water (aka curvature). This is time consuming to calculate for large molecules, however once the calculations are performed for two such, the differential occlusion between them is easier to calculate. In short we hope to have an immediate estimate of the buried area as two molecules are moved relative to each other. The final energy term, desolvation, is the hardest to calculate precisely. Typically it involves numerous DelPhi calculations each of which takes up to a minute on a low­end Iris. However, there are certain short cuts which can be taken when the problem is to calculate the relative change of solvation for the arrangements of two otherwise static molecules. These revolve around reformulating desolvation as a surface term akin to accessible area.

For both the hydrophobic and the desolvation penalty the approach that will be taken is to "prepare" certain sets of data for each molecule before attempting to "dock" them. This approach will also be taken to improve the speed of calculating the Van der Waals and Coulombic terms. This is especially important in the case of the Coulombic term because of the long range nature of the force precludes continuous direct (i.e. exact) calculation except where a small number of charges are involved (i.e. typically a hundred for each subset).

The resultant energies will only be "approximate" in that they will not exactly reproduce the numbers that would be obtained from a precise application of each theoretical model. But since we should remember that each theoretical model term is only a model of physical reality the more important question is as to whether these approximations will still accurately reflect enough of the underlying physics to be useful. For instance, providing a sufficiently large penalty for burying two oppositely charged groups together compared to the favorable Coulombic energy. This will probably only be truly tested by extensive use.

The energies calculated could be displayed both numerically and graphically as the molecules are arranged. In some cases where the contributions can be made local (e.g. Coulombic) these can be displayed as an atom or surface property.

Using reduced surface representations it might be possible to automatically dock molecules together based upon their shape and local properties. One such promising method is to use geometric hashing to check all such matches. Such approaches will be investigated.

Appendix A - File Formats

A.1 - Grasp Surface File

This unformatted file starts with five lines of 80 characters each. Line 1 contains the words "format=1" (there are no other formats at this time). Line 2 contains key words for the information contained within: "vertices" for vertex positions, "accessibles" for associated accessible surface point coordinates, "normals" for the normal vector (of length unity) for each vertex, and "triangles" for the triangle index list (which is a list of integers such that entries i­2, i­1 and i, where mod(i,3)=0, give the vertices which make up triangle i/3). Line 3 contains keywords for variables also written to this file: "potentials", "curvature", "distances", "gproperty1", and "gproperty2". Line 4 contains the number of vertices, the number of triangles, the grid size of the lattice used to create the surface (the number of points along one edge of the cube, always 65), and the reciprocal lattice spacing. Line 5 contains the midpoint of the coordinate system from which the vertices were derived (the midpoint of the Grasp box).

The data then follows in the order of the keywords. These are all REAL*4 except for the triangle indices which are INTEGER*2.

A.2 - DelPhi Potential Map

This file is unformatted. Its contents follow the order:

character*20 uplbl

character*10 nxtlbl, character*60 toplbl

real*4 phi(65,65,65)

character*16 botlbl

real*4 scale, mid(3)

Here phi contains the map information, scale the reciprocal grid spacing, and mid the grid midpoint. The rest are just character strings containing non­Grasp information.

A.3 - DelPhi Charge File

This file can have any number of comment lines in the header. After the last comment line should appear the following "keyword" line


Any comments after this line must be to the right of character 22, since this part of the line is not interpreted as an assignment.

To make an assignment statement, the user places the atom name specification under the four characters "atom" in the above line, the three characters of the residue name under "res", the residue number under "numb", the chain designator under "c", and the charge value under "charge". If a descriptor field left totally blank it is treated as wild, so if the chain specifier is blank the assignments spelled out by the other fields in that line are applied to all chains in the molecule. However, blank spaces within a field are treated as true blanks, so "c " in the atom field will not apply to a "ca " atom. This is different from how DelPhi radii files are dealt with.

Individual assignment statements are interpreted identically in both Grasp and DelPhi. However, the method of eventual assignment differs. In DelPhi each such specification is entered in a hash table and after all lines are read, the charge assigned to a particular atom is that which is most specifically declared in the assignment statements. On the other hand, the method Grasp uses to assign charges from a DelPhi charge file is that each line is interpreted individually before the next is read.

As an example of the different approaches, the file


nz lys 25 A 0.0

nz lys 1.0

will charge all zeta nitrogens of the molecule in Grasp, and all BUT lysine 25 on chain A in DelPhi. If the line order were reversed, the charging would be identical since Grasp would change the charge on lysine 25 on chain A back to zero after first setting it to 1.0. Hence when using DelPhi charge files in Grasp be sure that the more general charge assignments precede the more specific.

A.4 - DelPhi Radius File

Radius assignment files in DelPhi are similar to, but simpler than, charge assignment files. There may comment lines before the initial "key word" line, which in this case is


Size files only contain room for atom name, residue name, and radius.

There is one further difference between DelPhi charge files and radius files, namely that in size files empty spaces within a descriptor field are treated as wild cards. Thus "c " under the "atom" header in a size file applies to all atoms whose names begin with "c".

The same difference in mode of assignment between DelPhi and Grasp occurs as is described above for charge files - the user should be careful to put more general assignments first if the same assignment is to be made by both programs.

Both charge and size files are supported by Grasp for DelPhi users' convenience. However, the Grasp user might want to consider using neither and instead using Grasp "alt" commands in a Grasp script file to assign radii and charges.

A.5 - Protein Data Bank (PDB) File and Grasp Variants

The complete specification for these files is quite complex. Grasp only uses information on lines beginning with "ATOM", "HETATM", or "TER". These lines are expected to have the Fortran format (a6, 5x, a5, 1x, a3, 1x, a1, a4, 4x, 3f8.3) in the first 54 characters (columns), where a6 is "ATOM " or "HETATM", a5 is the atom name (only four of which are used in Grasp), a3 the residue name, a1 the chain name, a4 the residue number, and 3f8.3 the xyz atom coordinates in Angstroms.

In some files columns 55-80 are also used to store information. In standard PDB format columns 55­60 are used for occupancy (f6.2) and columns 61­67 for B­value (f7.3). Grasp uses these fields as follows.

If the first two lines of the file are



columns 55-60 will be read as the radius and 61-67 as the charge of the atom.

If the first two lines of the file are



columns 55-60 will be read as general property 1 and 61-67 as general property 2 of the atom.

If the first two lines of the file are



columns 55-80 are read in free format as general properties 1 and 2 of the atom. This allows the user to store values to much higher accuracy.

Note that in using any of these formats there MUST be two numbers in some format in this space or an error may occur.

"TER" lines should occur after each molecule in a PDB file for Grasp to treat each as an independent molecule.

A.6 - Grasp Property File

This file contains a listing of a single property for either all atoms or all surface vertices. The user include as many comment lines as desired before a line which has the key word "surface=" or "atoms=" (so be careful not to include these in comment lines). All lines after that are assumed by the program to be numerical values (in any format). On the same line as the above key words the user should have one of the following: "potential", "distance", "curvature", "gproperty1", "gproperty2", "accessible", or "charge". For example, "surface=distance" will inform the program that the list of values should be placed in the distance array for surface vertices.

A.7 - Grasp Script File

This is a file where the user can store commands to be executed together, for instance a set of coloring commands for atoms. The extension for a script file is usually ".his" or ".gs". The first line must be "GRASP HISTORY FILE". Commands can be any valid command as defined in section 5. Any line beginning with an exclamation mark (!) is considered a comment. All other lines are sent to the command interpreter to be acted upon. There are examples of Grasp script files in the directory $GRASP/example_scripts_and_macros.

A script file can be written by Grasp. This file is a history file of all textport commands entered during a session. Note this does not include textport responses to menu-driven operations. This is a good place to start when constructing a script file, or it can be used to replay a session if the system crashed or just to get back to a particular situation.

Scripts are really just history replays and as such they expect a certain state of affairs to exist when they are started. If things are not as expected, a script will not perform properly. An example would be a script to create a molecular surface. If a surface has already been created, the program will normally prompt the user as to whether to replace the previous surface. If this was not a surface present when the script was created, it will not supply the answer to this question and so will get out of sequence. Therefore scripts should typically be run with a blank slate, i.e. immediately after starting the program. This does not apply to ".gs" files which are too simple to crash.

A.8 - Grasp Macro File

This a a file which contains a collection of commands which are given a name. Macro files generally are given the extension ".mac". The $GRASP directory contains the file default_macros which contains simple examples to do such things as surface a molecule or make a backbone worm. These can be used as models to create your own macros. There are other macro examples in the directory $GRASP/scripts_and_macros.

A macro can be run by selecting it on the Macros. A macro cannot be run unless its file has been read in. This is done with "Read: Grasp Macro File" and the user is prompted for a file name. The macros defined in the file will then appear by their assigned name on the Macros menu and selecting one will execute it.

Macros are designed to be run often and in many different circumstances. Therefore they are not as dependent on prgram state as scripts. If a macro encounters an unusual situation, it will query the user on how to proceed.

Some macros or scripts may expect entries from a file as opposed to the command line. Be sure to have such files if needed. If files are not found, the user is typically prompted for input.

A.9 - Pair-Wise Interaction (Matrix) Energy File

This file should have as its first line the words "GRASP RESIDUE INTERACTION". The next line should either be




The first format is more complete and expects lines in Fortran format (a3, 3x, i3, 10x, a3, 3x, i3, 10x, 12g). Here a3 is the first residue name, i3 the residue number, a3 the next residue name, i3 the next residue number, and 12g the interaction energy.

The second format contains only the residue numbers and the interaction strength (i5, 1x, i5, 4x, 12g).

There can be at most 10,000 lines of such descriptions. There is no provision for comments. Reading in a file automatically replaces all previous interaction values.

Appendix B - .init_Grasp Commands

One can set certain initial parameters for Grasp in a file named .init_Grasp. This file can be in the directory the user launches Grasp from, the user's home directory, or the data directory pointed to in the environment variable $GRASP. Commands have precedence in the same order - local over home over data directory. The commands are not case or spacing sensitive but they must contain the exact specifier words as listed below to the left of an equals sign, and similarly for the words or values to the right of the equals sign. Any line beginning with a "!" or a "#" is ignored as a comment. Lines which can not be interpreted as comments or commands are written to the textport as errors.

INITIAL DISPLAY=BONDS Set default molecule display to bonds

INITIAL DISPLAY=ATOMS Set default molecule display to atoms




DEFAULT ATOM DISPLAY=GPROPERTY1 Color atoms by general property 1

DEFAULT ATOM DISPLAY=GPROPERTY2 Color atoms by general property 2

DEFAULT ATOM DISPLAY=DISCRETE Color atoms via command line




DEFAULT SURFACE DISPLAY=GPROPERTY1 Color surface by general property 1

DEFAULT SURFACE DISPLAY=GPROPERTY2 Color surface by general property 2

DEFAULT SURFACE DISPLAY=DISCRETE Color surface via command line

DEFAULT SURFACE DISPLAY=LIT Surface is rendered and lit by SGI calls

DEFAULT SURFACE DISPLAY=PSEUDO Surface is rendered and lit by Grasp calls

DEFAULT SURFACE DISPLAY=MESH Surface is displayed as mesh

DEFAULT SURFACE DISPLAY=POINTS Surface is displayed as dots

Appendix C - Grasp Data Files

Grasp comes with certain data files which need to be installed in a known data directory.

The following two files are vital and Grasp will not function correctly without them:

default.siz is a DelPhi control file which assigns radii when a pdb file is read by the program. It may be edited by the user to include more specific radii assignments, such as different radii for different carbons, or to alter the radius values therein, for example setting hydrogen radii to zero. The format of these specifications is described in Appendix A.

v3.dat is an unformatted file which contains information necessary to the program to perform marching cubes surface construction.

The following five DelPhi charge files are non­essential, but they should nevertheless be installed for ease of use:

full.crg describes the charge assignments to each normally ionizable residue, i.e. aspartate, glutamate, lysine, arginine. (Histidines must be charged by the user.). This is a default charge set for most proteins.

full+backbone.crg adds charges to the atoms N, HN, CA, C and O of the backbone. These "partial" charges are taken from the CHARMM charge set. If no hydrogens are present, the user should use first full.crg, then back_no_h.crg.

back_no_h.crg compresses the hydrogen charge onto the nitrogen.

amber.crg contains assignments from the AMBER charge set developed by Kolmann, which is a complete partial charge set which will assign charges to nearly all atoms.

dnarna.crg contains partial charges for DNA and RNA taken from Tung, Harvey and McCammon, 1984, Biopolymers 23, 2173­2193.

Note that because of the variability of atom names in pdb files few guarantees can be made that the protein or DNA will correctly charge. The user should read the section on charging molecules for ways to check this procedure.

Remember that reading charge files does not necessarily wipe out old assignments, i.e. one should not attempt to change charge sets merely by reading in a different charge set without being sure that all previous assignments are overwritten. The user can set all charges to zero prior to reading a new charge file with the command "alt(q=0.0)".

The following two files are also included:

defcol.dat contains a set of default colors the user may wish to use. This will be read in upon program startup if it is in the current directory.

atom.col is a Grasp Script which will color atoms by a certain color scheme. This has to be read in via the menu selections "Read: Grasp Script File".

Both of the above files can be edited to the user's satisfaction.


Nicholls, A., Sharp, K. A., and Honig, B. (1991) Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins Struct. Funct. Gen. 11, 282.

W.Kabsch (1976) Acta Cryst. A32, 922.

Bonds as Sticks
Kuznetsov, D. A., and Lim, H. A. (1992) VisiCoor: a simple program for visualization of proteins. J. Mol. Graphics 10, 25-28.

Small Molecule Solvation
Jean-Charles, A., Nicholls, A., Sharp, K., Honig, B., Tempczyk, A., Hendrickson, T. H., and Still, W. C. (1991) Electrostatic contributions to solvation energies: comparisons of free energy perturbation and continuum calculations. J. Am. Chem. Soc. 113, 1454.

pKa Shifts
Yang, A.-S., Gunner, M. R., Sampogna, R., Sharp, K., and Honig, B. (1993) On the Calculations of pKas in Proteins. Proteins 15, 252­265.

Site­Site Interactions
Friedman, R. A., and Honig, B. (1992) The electrostatic contributions to DNA base­stacking interactions. Biopolymers 32, 145­159.

Redox Potentials
Gunner, M. R., and Honig, B. (1991) Electrostatic control of midpoint potentials in the cytochrome subunit of the Rhodopseudomonas viridis reaction center. Proc. Natl. Acad. Sci. 88, 9151­9155.

Potentials and Fields Around a Protein
Sharp, K, Fine, R., and Honig, B. (1987) Computer simulations of the diffusion of a substrate to an active site of an enzyme. Science 236, 1460­1463.

The DelPhi Algorithm
Nicholls, A., and Honig, B. (1991) A rapid finite difference algorithm, utilizing successive over­relaxation to solve the Poisson­Boltzmann equation. J. Comp. Chem. 12, 435­445.