1. Specifying a force field Amber is designed to work with several simple types of force field, although it is most commonly used with parame- terizations developed by Peter Kollman and his co-workers. One significant recent development is that there are now a variety of such parameterizations, with no obvious "default" value. The "traditional" parameterization uses fixed par- tial charges, centered on atoms. Examples of this are ff86, ff94, ff96, ff98 and ff99 (described below). The default in versions 5 and 6 of Amber was ff94; a comparable default now would probably be ff99, but users should consult the papers listed below to see a detailed discussion of the changes made. Less extensively used, but very promising, recent modi- fications add polarizable dipoles to atoms, so that the charge description depends upon the environment; such poten- tials are called "polarizable" or "non-additive". Examples are ff02 and ff02EP: the former has atom-based charges (as in the traditional parameterization), and the latter adds in off-center charges (or "extra points"), primarily to help describe better the angular dependence of hydrogen bonds. Again, users should consult the papers cited below to see details of how these new force fields have been developed. In order to tell LEaP which force field is being used, the four types of information described below need to be provided. This is generally accomplished by selecting an appropriate leaprc file, which loads the information needed for a specific force field. (See section 2.2, below). (1) A listing of the atom types, what elements they cor- respond to, and their hybridizations. This informa- tion is encoded as a set of LEaP commands, and is normally read from a leaprc file. (2) Residue descriptions (or "topologies") that describe the chemical nature of amino acids, nucleotides, and so on. These files specify the connectivities, atom types, charges, and other information. These files have a "prep" format (a now-obsolete part of Amber) and have a ".in" extension. Standard libraries of residue descriptions are in the amber7/dat/leap/prep directory. The antechamber program may be used to generate prep files for other organic molecules. (3) Parameter files give force constants, equilibrium bond lengths and angles, Lennard-Jones parameters, and the like. Standard files have a ".dat" exten- sion, and are found in amber6/dat/leap/parm. (4) Extensions or changes to the parameters can be included in frcmod files. The expectation is that the user will load a large, "standard" parameter file, and (if needed) a smaller frcmod file that keeps track of any changes to the default parameters that are needed. The frcmod files for changing the default water model (which is TIP3P) into other water models are in files like amber7/dat/leap/parm/frc- mod.tip4p. The parmchk program (part of Antechamber) can also generate frcmod files. 1.1. Description of the database files The following files are in the amber7/dat/leap direc- tory. Files with a ".in" extension are in the prep subdi- rectory, those with a ".dat" extension are in the parm sub- directory, as are the "frcmod" files. ____________________________________________________________ Amber 1999 and 2002 force fields parm99.dat Force field, for amino acids and some organic molecules; can be used with either additive or non-additive treatment of electrostatics parm99EP.dat Like parm99.dat, but with "extra-points": off-center atomic charges, somewhat like lone-pairs gaff.dat Newer (and still experimental) force field for quite general organic molecules. all_nuc02.in Nucleic acid input for building database, for a non- additive (polarizable) force field without extra points. all_amino02.in Amino acid input ... all_aminoct02.in COO- amino acid input ... all_aminont02.in NH3+ amino acid input .... all_nuc02EP.in Nucleic acid input for building database, for a non- additive (polarizable) force field with extra points. all_amino02EP.in Amino acid input ... all_aminoct02EP.in COO- amino acid input ... all_aminont02EP.in NH3+ amino acid input .... Amber 1994 (Cornell et al.) force field all_nuc94.in Nucleic acid input for building database. all_amino94.in Amino acid input for building database. all_aminoct94.in COO- amino acid input for database. all_aminont94.in NH3+ amino acid input for database. nacl.in Ion file parm94.dat 1994 force field file. parm96.dat modified version of 1994 force field, for proteins parm98.dat modified version of 1994 force field, for nucleic acids Amber 1984, 1986 (Weiner et al.) force fields all.in All atom database input. allct.in All atom database input, COO- Amino acids. allnt.in All atom database input, NH3+ Amino acids. uni.in United atom database input. unict.in United atom database input, COO- Amino acids. unint.in United atom database input, NH3+ Amino acids. parm91X.dat Parameters for 1984, 1986 force fields Solvent models: water.in topology definition for several water models meoh.in topology file for methanol chcl3.in topology file for chloroform nma.in topology file for N-methylacetamide frcmod.tip4p parameter changes from TIP3P -> TIP4P frcmod.tip5p parameter changes from TIP3P -> TIP5P frcmod.spce parameter changes from TIP3P -> SPC/E frcmod.pol3 parameter changes from TIP3P -> POL3 frcmod.meoh paramters for methanol frcmod.chcl3 paramters for chloroform frcmod.nma paramters for N-methyacetamide Miscellaneous: nucgen.dat Nucgen nucleic acid conformations. PROTON_INFO* Files needed for protonate map.DG-AMBER needed for NMR input generation. ____________________________________________________________ 1.2. Specifying which force field you want in LEaP Various combinations of the above files make sense, and we have moved to an "ff" (force field) nomenclature to iden- tify these; examples would then be ff94 (which was the default in Amber 5 and 6), ff99, etc. The most straightfor- ward way to specify which force field you want is to use one of the leaprc files in $AMBERHOME/dat/leap/cmd. The sytax is: xleap -s -f Here, the -s flag tells LEaP to ignore any leaprc file it might find, and the -f flag tells it to start with commands for some other file. Here are the combinations we support and recommend: +-------------------------------------------------------------+ | How to specify force fields in LEaP | | filename topology parameters | +-------------------------------------------------------------+ |leaprc.ff86 Weiner et al. 1986 parm91X.dat | |leaprc.ff94 Cornell et al. 1994 parm94.dat | |leaprc.ff96 " parm96.dat | |leaprc.ff98 " parm98.dat | |leaprc.ff99 " parm99.dat | |leaprc.ff02 reduced (polarizable) charges parm99.dat | |leaprc.ff02EP " + extra points parm99EP.dat | |leaprc.gaff none gaff.dat | +-------------------------------------------------------------+ Notes: (1) Unlike previous versions of Amber, there is no default leaprc file anymore. If you make a link from one of the files above to a file named leaprc, then that will become the default. For example: cd $AMBERHOME/dat/leap/cmd ln -s leaprc.ff99 leaprc will provide a good default for many users; after this you could just invoke tleap or xleap without any arguments, and it would automatically load the ff99 force field. If you put leaprc.ff94 in the above link command, you would be making the Cornell et al. force field the default, which was the behavior of versions 5 and 6 of Amber. Note also that a leaprc file in the current directory overrides any other such files that might be present in the search path. (2) The first five choices in the above table are for additive (non-polarizable) simulations; you should use saveAmberParm (or saveAmberParmPert) to save the prmtop file, and keep the default ipol=0 in sander or gibbs. (3) The ff02 entries in the above table are for non-addi- tive (polarizable) force fields. Use saveAmberParm- Pol to save the prmtop file, and set ipol=1 in the sander or gibbs input file. Note that POL3 is a polarizable water model, so you need to use saveAm- berParmPol for it as well. (4) The files above assume that nucleic acids are DNA, if not explicitly specified. Use the files leaprc.rna.ff98, leaprc.rna.ff99, leaprc.rna.ff02 or leaprc.rna.ff00EP to make the default RNA. If you have mixture of DNA and RNA, you will need to edit your PDB file, or use the loadPdbUsingSequence command in LEaP (see that chapter) in order to spec- ify which nucelotide is which. (5) There is also a leaprc.gaff file, which sets you up for the "general" Amber force field. This is primar- ily for use with Antechamber (see that chapter), and does not load any topology files. (6) The leaprc.ff86 files gives the 1986 all-atom parame- ters; Amber no longer directly supports the 1984 united atom parameter set. (7) Our experience with generalized Born simulations is all with ff94 or ff99; the current GB models are not compatible with polarizable force fields. The GB options igb=3 or 4 (see Chapter 5) were derived for use with ff94. Replacing explicit water with a GB model is equivalent to specifying a different force field, and users should be aware that none of the GB options (in Amber or elsewhere) is as "mature" as simulations with explicit solvent; user discretion is advised! 1.3. 1999 and 2002 force fields The ff99 force field [1] represents a new direction for Amber-related force fields, pointing towards "general" organic and bioorganic systems. The atom types are mostly those of Cornell et al. (see below), but changes have been made in many torsional parameters, and this parameterization supports both additive and non-additive (polarizable) force fields. The topology and coordinate files for the small molecule test cases used in the development of this force field are in the parm99_lib subdirectory. The ff99 force field uses these parameters, along with the topologies and charges from the Cornell et al. force field, to create an all-atom nonpolarizable force field for proteins and nucleic acids. This is probably the best "general purpose" force field included here for biomolecules. The ff02 force field is a polarizable variant of ff99. Here, the charges were determined at the B3LYP/cc- pVTZ//HF/6-31g* level, and hence are more like "gas-phase" charges. During charge fitting the correction for intramolecular self polarization has been included [2]. Bond polarization arising from interactions with a condensed phase environment are achieved through polarizable dipoles attached to the atoms. These are determined from isotropic atomic polarizabilities assigned to each atom, taken from experimental work of Applequist. The dipoles can either be determined at each step through an iterative scheme, or can be treated as additional dynamical variables, and propagated through dynamics along with the atomic positions, in a man- ner analogous Car-Parinello dynamics. Derivation of the polarizable force field required only minor changes in dihe- dral terms and a few modification of the van der Waals parameters. The user also has a choice to use the polarizable force field with extra points on which additional point charges are located; this is called ff02EP. The additional points are located on electron donating atoms (e.g. O,N,S), which mimic the presence of electron lone pairs [3]. For nucleic acids we chose to use extra interacting points only on nucleic acid bases and not on sugars or phosphate groups. There is not (yet) a full published description of this, but a good deal of preliminary work on small molecules is available [2,4]. Beyond small molecules, our intial tests have focussed on small proteins and double helical oligonucleotides, in additive TIP3P water solution. Such a simulation model, (using a polarizable solute in a non- polarizable solvent) gains some of the advantages of polar- ization at only a small extra cost, compared to a standard force field model. In particular, the polarizable force field appears better suited to reproduce intermolecular interactions and directionality of H-bonding in biological systems than the additive force field. Initial tests show ff02EP behaves slightly better than ff02, but it is not yet clear how significant or widespread these differences will be. The gaff.dat ("general Amber force field") is yet a further step towards general purpose organic molecules. It is primarily used in conjunction with the antechamber pro- gram, and users should consult that chapter for more infor- mation. A paper describing these parameters is being pre- pared for publication. 1.4. The Cornell et al. (1994) force field Contained in ff94 are parameters from the so-called "second generation" force field developed in the Kollman group in the early 1990's [5]. These parameters are espe- cially derived for solvated systems, and when used with an appropriate 1-4 electrostatic scale factor, have been shown to perform well at modelling many organic molecules. The parameters in parm94.dat omit the hydrogen bonding terms of earlier force fields. This is an all-atom force field; no united-atom counterpart is provided. 1-4 electrostatic interactions are scaled by 1.2 instead of the value of 2.0 that had been used in earlier force fields. Charges were derived using Hartree-Fock theory with the 6-31G* basis set, because this exaggerates the dipole moment of most residues by 10-20%. It thus "builds in" the amount of polarization which would be expected in aqueous solution. This is necessary for carrying out condensed phase simula- tions with an effective two-body force field which does not include explicit polarization. The charge-fitting procedure is described in Appendix D. The ff96 force field [6] differs from parm94.dat in that the torsions for and have been modified in response to ab initio calculations [7] which showed that the energy difference between conformations were quite different than calculated by Cornell et al. (using parm94.dat). To create parm96.dat, common V1 and V2 parameters were used for and , which were empirically adjusted to reproduce the energy dif- ference between extended and constrained alpha helical ener- gies for the alanine tetrapeptide. This led to a signifi- cant improvement between molecular mechanical and quantum mechanical relative energies for the remaining members of the set of tetrapeptides studied by Beachy et al. Users should be aware that parm96.dat has not been as extensively used as parm94.dat, and that it almost certainly has its own biases and idiosyncracies [8,9]. The ff98 force field [10] differs from parm94.dat in torsion angle parameters involving the glycosidic torsion in nucleic acids. These serve to improve the predicted helical repeat and sugar pucker profiles. 1.5. The Weiner et al. (1984,1986) force fields The ff86 parameters are described in early papers from the Kollman and Case groups [11,12]. [The "parm91" designa- tion is somewhat unfortunate: this file is really only a corrected version of the paramters described in the 1984 and 1986 papers listed above.] These parameters are not gener- ally recommended any more, but may still be useful for vac- uum simulations of nucleic acids and proteins using a dis- tance-dependent dielectric, or for comparisons to earlier work. The material in parm91X.dat is the parameter set dis- tributed with Amber 4.0. The STUB nonbonded set has been copied from parmuni.dat; these sets of parameters are appro- priate for united atom calculations using the "larger" car- bon radii referred to in the "note added in proof" of the 1984 JACS paper. If these values are used for a united atom calculation, the parameter scnb should be set to 8.0; for all-atom calculations use 2.0. The scee parameter should be set to 2.0 for both united atom and all-atom variants. Note that the default value for scee is sander is now 1.2 (the value for 1994 and later force fields; users must explicitly change this in their inputs for the earlier force fields. A number of terms in the non-bonded list of parm91X.dat should be noted. The non-bonded terms for I(iodine),CU(cop- per) and MG(magnesium) have not been carefully calibrated, but are given as approximate values. In the STUB set of non-bonded parameters, we have included parameters for a large hydrated monovalent cation (IP) that represent work by Singh et al 1985 on large hydrated counterions for DNA. Sim- ilar values are included for a hydrated anion (IM). The non-bonded potentials for hydrogen-bond pairs in ff86 uses a Lennard-Jones 10-12 potential. If you want to run sander with ff86, you will need to recompile, adding the -DHAS_10_12 flag to your MACHINE file. 1.6. Ions For alkali ions with TIP3P waters, we have provided the values of Aqvist [13], which are adjusted for Amber's non- bonded atom pair combining rules to give the same ion-OW potentials as in the original (which were designed for SPC water); these values reproduce the first peak of the radial distribution for ion-OW and the relative free energies of solvation in water of the various ions. Note that these values would have to be changed if a water model other than TIP3P were to be used. These potentials may also need modi- fication if absolute free energies of solvation are impor- tant [14]. 1.7. Solvent models Amber now provides direct support for several water models. The default water model is TIP3P [15]. This model will be used for residues with names HOH or WAT. If you want to use other water models, execute the following leap commands after loading your leaprc file: WAT = PL3 (residues named WAT in pdb file will be POL3) loadAmberParams frcmod.pol3 (sets the HW,OW parameters to POL3) (The above is obviously for the POL3 model.) The water.lib file contains TIP3P [15], TIP4P [15,16], TIP5P [17], POL3 [18] and SPC/E [19] models for water; these are called TP3, T4P, T5P, PL3 and SPC, respectively. By default, the residue name in the prmtop file will be WAT, regardless of which water model is used. If you want to change this (in order to keep track of which water model you are using, say), you can change the residue name to whatever you like. For example, WAT = TP4 set WAT.1 name "TP4" would make a special label in PDB and prtmop files for TIP4P water. Note that Brookhaven format files allow at most three characters for the residue label. In addition, non-polarizable models for the organic solvents methanol, chloroform and N-methylacetamide are pro- vided. The input files for a single molecule are in amber7/dat/leap/prep, and the corresponding frcmod files are in amber7/dat/leap/parm. Pre-equlibrated boxes are in amber7/dat/leap/lib. For example, to solvate a simple pep- tide in methanol, you could do the following: source leaprc.ff99 (get a standard force field) loadAmberParams frcmod.meoh (get methanol parameters) peptide = sequence { ACE VAL NME } (construct a simple peptide) solvateBox peptide MEOHBOX 12.0 0.8 (solvate the peptide with meoh) saveAmberParm peptide prmtop prmcrd quit Similar commands will work for other solvent models. 1. J. Wang, P. Cieplak & P.A. Kollman. How well does a restrained electrostatic potential (RESP) model perform in calcluating conformational energies of organic and biological molecules?. J. Comput. Chem. 21, 1049-1074(2000). 2. P. Cieplak, J. Caldwell & P. Kollman. Molecular Mechan- ical Models for Organic and Biological Systems Going Beyond the Atom Centered Two Body Additive Approxima- tion: Aqueous Solution Free Energies of Methanol and N- Methyl Acetamide, Nucleic Acid Base, and Amide Hydrogen Bonding and Chloroform/Water Partition Coefficients of the Nucleic Acid Bases. J. Computat. Chem. 22, 1048-1057(2001). 3. R.W. Dixon & P.A. Kollman. Advancing Beyond the Atom- Centered Model in Additive and Nonadditive Molecular Mechanics.. J. Computat. Chem. 18, 1632-1646(1997). 4. E. Meng, P. Cieplak, J.W. Caldwell & P.A. Kollman. Accurate solvation free energies of acetate and methylammonium ions calculated with a polarizable water model. J. Am. Chem. Soc. 116, 12061-12062(1994). 5. W.D. Cornell, P. Cieplak, C.I. Bayly, I.R. Gould, K.M. Merz, Jr., D.M. Ferguson, D.C. Spellmeyer, T. Fox, J.W. Caldwell & P.A. Kollman. A second generation force field for the simulation of proteins, nucleic acids, and organic molecules. J. Am. Chem. Soc. 117, 5179-5197(1995). 6. P.A. Kollman, R. Dixon, W. Cornell, T. Fox, C. Chipot & A. Pohorille. The development/application of a 'mini- malist' organic/biochemical molecular mechanic force field using a combination of ab initio calculations and experimental data. In Computer Simulation of Biomolec- ular Systems, Vol. 3, A. Wilkinson, P. Weiner & W.F. van Gunsteren, Ed. Elsevier, (1997). pp. 83-96. 7. M.D. Beachy & R.A. Friesner. J. Am. Chem. Soc. 119, 5908-5920(1997). 8. L. Wang, Y. Duan, R. Shortle, B. Imperiali & P.A. Kollman. Study of the stability and unfolding mechanism of BBA1 by molecular dynamics simulations at different termperatures. Prot. Sci. 8, 1292-1304(1999). 9. J. Higo, N. Ito, M. Kuroda, S. Ono, N. Nakajima & H. Nakamura. Energy landscape of a peptide consisting of -helix, 310 helix, -turn, -hairpin and other disordered conformations. Prot. Sci. 10, 1160-1171(2001). 10. T.E. Cheatham, III, P. Cieplak & P.A. Kollman. A modi- fied version of the Cornell et al. force field with improved sugar pucker phases and helical repeat. J. Biomol. Struct. Dyn. 16, 845-862(1999). 11. S.J. Weiner, P.A. Kollman, D.A. Case, U.C. Singh, C. Ghio, G. Alagona, S. Profeta, Jr. & P. Weiner. A new force field for molecular mechanical simulation of nucleic acids and proteins. J. Am. Chem. Soc. 106, 765-784(1984). 12. S.J. Weiner, P.A. Kollman, D.T. Nguyen & D.A. Case. An all-atom force field for simulations of proteins and nucleic acids. J. Computat. Chem. 7, 230-252(1986). 13. J. qvist. Ion-water interaction potentials derived from free energy perturbation simulations. J. Phys. Chem. 94, 8021-8024(1990). 14. T. Darden, D. Pearlman & L.G. Pedersen. Ionic charging free energies: Spherical versus periodic boundary con- ditions. J. Chem. Phys. 109, 10921-10935(1998). 15. W.L. Jorgensen, J. Chandrasekhar, J. Madura & M.L. Klein. Comparison of Simple Potential Functions for Simulating Liquid Water. J. Chem. Phys. 79, 926-935(1983). 16. W.L. Jorgensen & J.D. Madura. Mol. Phys. 56, 1381(1985). 17. M.W. Mahoney & W.L. Jorgensen. A five-site model for liquid water and the reproduction of the density anomaly by rigid, nonpolarizable potential functions. J. Chem. Phys. 112, 8910-8922(2000). 18. J.W. Caldwell & P.A. Kollman. Structure and properties of neat liquids using nonadditive molecular dynamics: Water, methanol and N-methylacetamide. J. Phys. Chem. 99, 6208-6219(1995). 19. H.J.C. Berendsen, J.R. Grigera & T.P. Straatsma. The missing term in effective pair potentials. J. Phys. Chem. 91, 6269-6271(1987).