pdb2gmx input files¶

The GROMACS program pdb2gmx generates a topology for the input coordinate file. Several formats are supported for that coordinate file, but pdb is the most commonly-used format (hence the name pdb2gmx). pdb2gmx searches for force fields in sub-directories of the GROMACS share/top directory and your working directory. Force fields are recognized from the file forcefield.itp in a directory with the extension .ff. The file forcefield.doc may be present, and if so, its first line will be used by pdb2gmx to present a short description to the user to help in choosing a force field. Otherwise, the user can choose a force field with the -ff xxx command-line argument to pdb2gmx, which indicates that a force field in a xxx.ff directory is desired. pdb2gmx will search first in the working directory, then in the GROMACS share/top directory, and use the first matching xxx.ff directory found.

Two general files are read by pdb2gmx: an atom type file (extension atp, see Atom types) from the force-field directory, and a file called residuetypes.dat from either the working directory, or the GROMACS share/top directory. residuetypes.dat determines which residue names are considered protein, DNA, RNA, water, and ions.

pdb2gmx can read one or multiple databases with topological information for different types of molecules. A set of files belonging to one database should have the same basename, preferably telling something about the type of molecules (e.g. aminoacids, rna, dna). The possible files are:

<basename>.rtp
<basename>.r2b (optional)
<basename>.arn (optional)
<basename>.hdb (optional)
<basename>.n.tdb (optional)
<basename>.c.tdb (optional)

Only the rtp file, which contains the topologies of the building blocks, is mandatory. Information from other files will only be used for building blocks that come from an rtp file with the same base name. The user can add building blocks to a force field by having additional files with the same base name in their working directory. By default, only extra building blocks can be defined, but calling pdb2gmx with the -rtpo option will allow building blocks in a local file to replace the default ones in the force field.

Residue database¶

The files holding the residue databases have the extension rtp. Originally this file contained building blocks (amino acids) for proteins, and is the GROMACS interpretation of the rt37c4.dat file of GROMOS. So the residue database file contains information (bonds, charges, charge groups, and improper dihedrals) for a frequently-used building block. It is better not to change this file because it is standard input for pdb2gmx, but if changes are needed make them in the top file (see Topology file), or in a rtp file in the working directory as explained in sec. pdb2gmx input files. Defining topologies of new small molecules is probably easier by writing an include topology file itp directly. This will be discussed in section Molecule.itp file. When adding a new protein residue to the database, do not forget to add the residue name to the residuetypes.dat file, so that grompp, make_ndx and analysis tools can recognize the residue as a protein residue (see Default Groups).

The rtp files are only used by pdb2gmx. As mentioned before, the only extra information this program needs from the rtp database is bonds, charges of atoms, charge groups, and improper dihedrals, because the rest is read from the coordinate input file. Some proteins contain residues that are not standard, but are listed in the coordinate file. You have to construct a building block for this “strange” residue, otherwise you will not obtain a top file. This also holds for molecules in the coordinate file such as ligands, polyatomic ions, crystallization co-solvents, etc. The residue database is constructed in the following way:

[ bondedtypes ]  ; mandatory
; bonds  angles  dihedrals  impropers
     1       1          1          2  ; mandatory

[ GLY ]  ; mandatory

 [ atoms ]  ; mandatory
; name  type  charge  chargegroup
     N     N  -0.280     0
     H     H   0.280     0
    CA   CH2   0.000     1
     C     C   0.380     2
     O     O  -0.380     2

 [ bonds ]  ; optional
;atom1 atom2      b0      kb
     N     H
     N    CA
    CA     C
     C     O
    -C     N

 [ exclusions ]  ; optional
;atom1 atom2

 [ angles ]  ; optional
;atom1 atom2 atom3    th0    cth

 [ dihedrals ]  ; optional
;atom1 atom2 atom3 atom4   phi0     cp   mult

 [ impropers ]  ; optional
;atom1 atom2 atom3 atom4     q0     cq
     N    -C    CA     H
    -C   -CA     N    -O

[ ZN ]

 [ atoms ]
    ZN    ZN   2.000     0

The file is free format; the only restriction is that there can be at most one entry on a line. The first field in the file is the [ bondedtypes ] field, which is followed by four numbers, indicating the interaction type for bonds, angles, dihedrals, and improper dihedrals. The file contains residue entries, which consist of atoms and (optionally) bonds, angles, dihedrals, and impropers. The charge group codes denote the charge group numbers. Atoms in the same charge group should always be ordered consecutively. When using the hydrogen database with pdb2gmx for adding missing hydrogens (see hdb), the atom names defined in the rtp entry should correspond exactly to the naming convention used in the hydrogen database. The atom names in the bonded interaction can be preceded by a minus or a plus, indicating that the atom is in the preceding or following residue respectively. Explicit parameters added to bonds, angles, dihedrals, and impropers override the standard parameters in the itp files. This should only be used in special cases. Instead of parameters, a string can be added for each bonded interaction. This is used in GROMOS-96 rtp files. These strings are copied to the topology file and can be replaced by force-field parameters by the C-preprocessor in grompp using #define statements.

pdb2gmx automatically generates all angles. This means that for most force fields the [ angles ] field is only useful for overriding itp parameters. For the GROMOS-96 force field the interaction number of all angles needs to be specified.

pdb2gmx automatically generates one proper dihedral for every rotatable bond, preferably on heavy atoms. When the [ dihedrals ] field is used, no other dihedrals will be generated for the bonds corresponding to the specified dihedrals. It is possible to put more than one dihedral function on a rotatable bond. In the case of CHARMM27 FF pdb2gmx can add correction maps to the dihedrals using the default -cmap option. Please refer to CHARMM for more information.

pdb2gmx sets the number of exclusions to 3, which means that interactions between atoms connected by at most 3 bonds are excluded. Pair interactions are generated for all pairs of atoms that are separated by 3 bonds (except pairs of hydrogens). When more interactions need to be excluded, or some pair interactions should not be generated, an [ exclusions ] field can be added, followed by pairs of atom names on separate lines. All non-bonded and pair interactions between these atoms will be excluded.

Residue to building block database¶

Each force field has its own naming convention for residues. Most residues have consistent naming, but some, especially those with different protonation states, can have many different names. The r2b files are used to convert standard residue names to the force-field build block names. If no r2b is present in the force-field directory or a residue is not listed, the building block name is assumed to be identical to the residue name. The r2b can contain 2 or 5 columns. The 2-column format has the residue name in the first column and the building block name in the second. The 5-column format has 3 additional columns with the building block for the residue occurring in the N-terminus, C-terminus and both termini at the same time (single residue molecule). This is useful for, for instance, the AMBER force fields. If one or more of the terminal versions are not present, a dash should be entered in the corresponding column.

There is a GROMACS naming convention for residues which is only apparent (except for the pdb2gmx code) through the r2b file and specbond.dat files. This convention is only of importance when you are adding residue types to an rtp file. The convention is listed in Table 12. For special bonds with, for instance, a heme group, the GROMACS naming convention is introduced through specbond.dat (see Special bonds), which can subsequently be translated by the r2b file, if required.

Table 12 Internal GROMACS residue naming convention.¶
GROMACS ID	Residue
ARG	protonated arginine
ARGN	neutral arginine
ASP	negatively charged aspartic acid
ASPH	neutral aspartic acid
CYS	neutral cysteine
CYS2	cysteine with sulfur bound to another cysteine or a heme
GLU	negatively charged glutamic acid
GLUH	neutral glutamic acid
HISD	neutral histidine with N\(_\delta\) protonated
HISE	neutral histidine with N\(_\epsilon\) protonated
HISH	positive histidine with both N\(_\delta\) and N\(_\epsilon\) protonated
HIS1	histidine bound to a heme
LYSN	neutral lysine
LYS	protonated lysine
HEME	heme

Atom renaming database¶

Force fields often use atom names that do not follow IUPAC or PDB convention. The arn database is used to translate the atom names in the coordinate file to the force-field names. Atoms that are not listed keep their names. The file has three columns: the building block name, the old atom name, and the new atom name, respectively. The residue name supports question-mark wildcards that match a single character.

An additional general atom renaming file called xlateat.dat is present in the share/top directory, which translates common non-standard atom names in the coordinate file to IUPAC/PDB convention. Thus, when writing force-field files, you can assume standard atom names and no further atom name translation is required, except for translating from standard atom names to the force-field ones.

Hydrogen database¶

The hydrogen database is stored in hdb files. It contains information for the pdb2gmx program on how to connect hydrogen atoms to existing atoms. In versions of the database before GROMACS 3.3, hydrogen atoms were named after the atom they are connected to: the first letter of the atom name was replaced by an ‘H.’ In the versions from 3.3 onwards, the H atom has to be listed explicitly, because the old behavior was protein-specific and hence could not be generalized to other molecules. If more than one hydrogen atom is connected to the same atom, a number will be added to the end of the hydrogen atom name. For example, adding two hydrogen atoms to ND2 (in asparagine), the hydrogen atoms will be named HD21 and HD22. This is important since atom naming in the rtp file must be the same. The format of the hydrogen database is as follows:

; res   # additions
        # H add type    H       i       j       k
ALA     1
        1       1       H       N       -C      CA
ARG     4
        1       2       H       N       CA      C
        1       1       HE      NE      CD      CZ
        2       3       HH1     NH1     CZ      NE
        2       3       HH2     NH2     CZ      NE

On the first line we see the residue name (ALA or ARG) and the number of kinds of hydrogen atoms that may be added to this residue by the hydrogen database. After that follows one line for each addition, on which we see:

The number of H atoms added
The method for adding H atoms, which can be any of:
1. one planar hydrogen, e.g. rings or peptide bond
  
  One hydrogen atom (n) is generated, lying in the plane of atoms (i,j,k) on the plane bisecting angle (j-i-k) at a distance of 0.1 nm from atom i, such that the angles (n-i-j) and (n-i-k) are \(>\) 90\(^{\rm o}\).
2. one single hydrogen, e.g. hydroxyl
  
  One hydrogen atom (n) is generated at a distance of 0.1 nm from atom i, such that angle (n-i-j)=109.5 degrees and dihedral (n-i-j-k)=trans.
3. two planar hydrogens, e.g. ethylene -C=CH\(_2\), or amide -C(=O)NH\(_2\)
  
  Two hydrogens (n1,n2) are generated at a distance of 0.1 nm from atom i, such that angle (n1-i-j)=(n2-i-j)=120 degrees and dihedral (n1-i-j-k)=cis and (n2-i-j-k)=trans, such that names are according to IUPAC standards 129.
4. two or three tetrahedral hydrogens, e.g. -CH\(_3\)
  
  Three (n1,n2,n3) or two (n1,n2) hydrogens are generated at a distance of 0.1 nm from atom i, such that angle (n1-i-j)=(n2-i-j)=(n3-i-j)=109.47\(^{\rm o}\), dihedral (n1-i-j-k)=trans, (n2-i-j-k)=trans+120 and (n3-i-j-k)=trans+240\(^{\rm o}\).
5. one tetrahedral hydrogen, e.g. C\(_3\)CH
  
  One hydrogen atom (n\(^\prime\)) is generated at a distance of 0.1 nm from atom i in tetrahedral conformation such that angle (n\(^\prime\)-i-j)=(n\(^\prime\)-i-k)=(n\(^\prime\)-i-l)=109.47\(^{\rm o}\).
6. two tetrahedral hydrogens, e.g. C-CH\(_2\)-C
  
  Two hydrogen atoms (n1,n2) are generated at a distance of 0.1 nm from atom i in tetrahedral conformation on the plane bisecting angle j-i-k with angle (n1-i-n2)=(n1-i-j)=(n1-i-k)=109.47\(^{\rm o}\).
7. two water hydrogens
  
  Two hydrogens are generated around atom i according to SPC 80 water geometry. The symmetry axis will alternate between three coordinate axes in both directions.
8. three water “hydrogens”
  
  Two hydrogens are generated around atom i according to SPC 80 water geometry. The symmetry axis will alternate between three coordinate axes in both directions. In addition, an extra particle is generated on the position of the oxygen with the first letter of the name replaced by ‘M’. This is for use with four-atom water models such as TIP4P 128.
9. four water “hydrogens”
  
  Same as above, except that two additional particles are generated on the position of the oxygen, with names ‘LP1’ and ‘LP2.’ This is for use with five-atom water models such as TIP5P 130.
The name of the new H atom (or its prefix, e.g. HD2 for the asparagine example given earlier).
Three or four control atoms (i,j,k,l), where the first always is the atom to which the H atoms are connected. The other two or three depend on the code selected. For water, there is only one control atom.

Some more exotic cases can be approximately constructed from the above tools, and with suitable use of energy minimization are good enough for beginning MD simulations. For example secondary amine hydrogen, nitrenyl hydrogen (\(\mathrm{C}=\mathrm{NH}\)) and even ethynyl hydrogen could be approximately constructed using method 2 above for hydroxyl hydrogen.

Termini database¶

The termini databases are stored in aminoacids.n.tdb and aminoacids.c.tdb for the N- and C-termini respectively. They contain information for the pdb2gmx program on how to connect new atoms to existing ones, which atoms should be removed or changed, and which bonded interactions should be added. Their format is as follows (from gromos43a1.ff/aminoacids.c.tdb):

[ None ]

[ COO- ]
[ replace ]
C   C       C       12.011  0.27
O   O1      OM      15.9994 -0.635
OXT O2      OM      15.9994 -0.635
[ add ]
2   8       O       C       CA      N
    OM      15.9994 -0.635
[ bonds ]
C   O1      gb_5
C   O2      gb_5
[ angles ]
O1  C       O2      ga_37
CA  C       O1      ga_21
CA  C       O2      ga_21
[ dihedrals ]
N   CA      C       O2      gd_20
[ impropers ]
C   CA      O2      O1      gi_1

The file is organized in blocks, each with a header specifying the name of the block. These blocks correspond to different types of termini that can be added to a molecule. In this example [ COO- ] is the first block, corresponding to changing the terminal carbon atom into a deprotonated carboxyl group. [ None ] is the second terminus type, corresponding to a terminus that leaves the molecule as it is. Block names cannot be any of the following: replace, add, delete, bonds, angles, dihedrals, impropers. Doing so would interfere with the parameters of the block, and would probably also be very confusing to human readers.

For each block the following options are present:

[ replace ]

Replace an existing atom by one with a different atom type, atom name, charge, and/or mass. This entry can be used to replace an atom that is present both in the input coordinates and in the rtp database, but also to only rename an atom in the input coordinates such that it matches the name in the force field. In the latter case, there should also be a corresponding [ add ] section present that gives instructions to add the same atom, such that the position in the sequence and the bonding is known. Such an atom can be present in the input coordinates and kept, or not present and constructed by pdb2gmx. For each atom to be replaced on line should be entered with the following fields:
- name of the atom to be replaced
- new atom name (optional)
- new atom type
- new mass
- new charge
[ add ]

Add new atoms. For each (group of) added atom(s), a two-line entry is necessary. The first line contains the same fields as an entry in the hydrogen database (name of the new atom, number of atoms, type of addition, control atoms, see hdb), but the possible types of addition are extended by two more, specifically for C-terminal additions:
1. two carboxyl oxygens, -COO\(^-\)
  
  Two oxygens (n1,n2) are generated according to rule 3, at a distance of 0.136 nm from atom i and an angle (n1-i-j)=(n2-i-j)=117 degrees
2. carboxyl oxygens and hydrogen, -COOH
  
  Two oxygens (n1,n2) are generated according to rule 3, at distances of 0.123 nm and 0.125 nm from atom i for n1 and n2, respectively, and angles (n1-i-j)=121 and (n2-i-j)=115 degrees. One hydrogen (n\(^\prime\)) is generated around n2 according to rule 2, where n-i-j and n-i-j-k should be read as n\(^\prime\)-n2-i and n\(^\prime\)-n2-i-j, respectively.
After this line, another line follows that specifies the details of the added atom(s), in the same way as for replacing atoms, i.e.:
- atom type
- mass
- charge
- charge group (optional)
Like in the hydrogen database (see rtp), when more than one atom is connected to an existing one, a number will be appended to the end of the atom name. Note that, like in the hydrogen database, the atom name is now on the same line as the control atoms, whereas it was at the beginning of the second line prior to GROMACS version 3.3. When the charge group field is left out, the added atom will have the same charge group number as the atom that it is bonded to.
[ delete ]

Delete existing atoms. One atom name per line.
[ bonds ], [ angles ], [ dihedrals ] and [ impropers ]

Add additional bonded parameters. The format is identical to that used in the rtp file, see rtp.

Virtual site database¶

Since we cannot rely on the positions of hydrogens in input files, we need a special input file to decide the geometries and parameters with which to add virtual site hydrogens. For more complex virtual site constructs (e.g. when entire aromatic side chains are made rigid) we also need information about the equilibrium bond lengths and angles for all atoms in the side chain. This information is specified in the vsd file for each force field. Just as for the termini, there is one such file for each class of residues in the rtp file.

The virtual site database is a simple list of information. The first couple of sections specify which mass centers (typically called MCH\(_3\)/MNH\(_3\)) to use for CH\(_3\), NH\(_3\), and NH\(_2\) groups. Depending on the equilibrium bond lengths and angles between the hydrogens and heavy atoms we need to apply slightly different constraint distances between these mass centers. Note that we do not have to specify the actual parameters (that is automatic), just the type of mass center to use. To accomplish this, there are three sections names [ CH3 ], [ NH3 ], and [ NH2 ]. For each of these we expect three columns. The first column is the atom type bound to the 2/3 hydrogens, the second column is the next heavy atom type which this is bound, and the third column the type of mass center to use. As a special case, in the [ NH2 ] section it is also possible to specify planar in the second column, which will use a different construction without mass center. There are currently different opinions in some force fields whether an NH\(_2\) group should be planar or not, but we try hard to stick to the default equilibrium parameters of the force field.

The second part of the virtual site database contains explicit equilibrium bond lengths and angles for pairs/triplets of atoms in aromatic side chains. These entries are currently read by specific routines in the virtual site generation code, so if you would like to extend it e.g. to nucleic acids you would also need to write new code there. These sections are named after the short amino acid names ([ PHE ], [ TYR ], [ TRP ], [ HID ], [ HIE ], [ HIP ]), and simply contain 2 or 3 columns with atom names, followed by a number specifying the bond length (in nm) or angle (in degrees). Note that these are approximations of the equilibrated geometry for the entire molecule, which might not be identical to the equilibrium value for a single bond/angle if the molecule is strained.

Special bonds¶

The primary mechanism used by pdb2gmx to generate inter-residue bonds relies on head-to-tail linking of backbone atoms in different residues to build a macromolecule. In some cases (e.g. disulfide bonds, a heme group, branched polymers), it is necessary to create inter-residue bonds that do not lie on the backbone. The file specbond.dat takes care of this function. It is necessary that the residues belong to the same [ moleculetype ]. The -merge and -chainsep functions of pdb2gmx can be useful when managing special inter-residue bonds between different chains.

The first line of specbond.dat indicates the number of entries that are in the file. If you add a new entry, be sure to increment this number. The remaining lines in the file provide the specifications for creating bonds. For these bonds, you can also optionally specify a custom improper dihedral associated with the new bond. The format of the lines, with optional entries in [], is as follows:

resA atomA nbondsA resB atomB nbondsB length newresA newresB [atomI atomJ atomK atomL]

The columns indicate:

resA The name of residue A that participates in the bond.
atomA The name of the atom in residue A that forms the bond.
nbondsA The total number of bonds atomA can form.
resB The name of residue B that participates in the bond.
atomB The name of the atom in residue B that forms the bond.
nbondsB The total number of bonds atomB can form.
length The reference length for the bond. If atomA and atomB are not within length \(\pm\) 10% in the coordinate file supplied to pdb2gmx, no bond will be formed.
newresA The new name of residue A, if necessary. Some force fields use e.g. CYS2 for a cysteine in a disulfide or heme linkage.
newresB The new name of residue B, likewise.
atomI Custom improper dihedral atom i of i-j-k-l. Has format
[specbond residue]-[atom name] (e.g. B-SG). The letter is either A or B corresponding to resA or resB, respectively.
atomJ Custom improper dihedral atom j of i-j-k-l.
atomK Custom improper dihedral atom k of i-j-k-l.
atomL Custom improper dihedral atom l of i-j-k-l.