Running membrane simulations in GROMACS

Running Membrane Simulations

Users frequently encounter problems when running simulations of lipid bilayers, especially when a protein is involved. Users seeking to simulate membrane proteins may find this tutorial useful.

One protocol for the simulation of membrane proteins consists of the following steps:

  1. Choose a force field for which you have parameters for the protein and lipids.
  2. Insert the protein into the membrane. (For instance, use g_membed on a pre-formed bilayer or do a coarse-grained self-assembly simulation and then convert back to the atomistic representation.)
  3. Solvate the system and add ions to neutralize excess charges and adjust the final ion concentration.
  4. Energy minimize.
  5. Let the membrane adjust to the protein. Typically run MD for ~5-10ns with restraints (1000 kJ/(mol nm2) on all protein heavy atoms.
  6. Equilibrate without restraints.
  7. Run production MD.

Adding waters with genbox

When generating waters around a pre-formed lipid membrane with solvate you may find that water molecules get introduced into interstices in the membrane. There are several approaches to removing these, including

  • a short MD run to get the hydrophobic effect to exclude these waters. In general this is sufficient to reach a water-free hydrophobic phase, as the molecules are usually expelled quickly and without disrupting the general structure. If your setup relies on a completely water-free hydrophobic phase at the start, you can try to follow the advice below:
  • Set the -radius option in gmx solvate to change the water exclusion radius,
  • copy vdwradii.dat from your $GMXLIB location to the working directory, and edit it to increase the radii of your lipid atoms (between 0.35 and 0.5nm is suggested for carbon) to prevent solvate from seeing interstices large enough for water insertion,
  • editing your structure by hand to delete them (remembering to adjust your atom count for gro files and to account for any changes in the topology), or
  • use a script someone wrote to remove them.

External material

Parameterization of novel molecules

Most of your parametrization questions/problems can be resolved very simply, by remembering the following two rules:

  • You should not mix and match force fields. Force fields are (at best) designed to be self-consistent, and will not typically work well with other force fields. If you simulate part of your system with one force field and another part with a different force field which is not parametrized with the first force field in mind, your results will probably be questionable, and hopefully reviewers will be concerned. Pick a force field. Use that force field.
  • If you need to develop new parameters, derive them in a manner consistent with how the rest of the force field was originally derived, which means that you will need to review the original literature. There isn’t a single right way to derive force field parameters; what you need is to derive parameters that are consistent with the rest of the force field. How you go about doing this depends on which force field you want to use. For example, with AMBER force fields, deriving parameters for a non-standard amino acid would probably involve doing a number of different quantum calculations, while deriving GROMOS or OPLS parameters might involve more (a) fitting various fluid and liquid-state properties, and (b) adjusting parameters based on experience/chemical intuition/analogy. Some suggestions for automated approaches can be found here.

It would be wise to have a reasonable amount of simulation experience with GROMACS before attempting to parametrize new force fields, or new molecules for existing force fields. These are expert topics, and not suitable for giving to (say) undergraduate students for a research project, unless you like expensive quasi-random number generators. A very thorough knowledge of Chapter 5 of the GROMACS Reference Manual will be required. If you haven’t been warned strongly enough, please read below about parametrization for exotic species.

Another bit of advice: Don’t be more haphazard in obtaining parameters than you would be buying fine jewellery. Just because the guy on the street offers to sell you a diamond necklace for $10 doesn’t mean that’s where you should buy one. Similarly, it isn’t necessarily the best strategy to just download parameters for your molecule of interest from the website of someone you’ve never heard of, especially if they don’t explain how they got the parameters.

Be forewarned about using PRODRG topologies without verifying their contents: the artifacts of doing so are now published, along with some tips for properly deriving parameters for the GROMOS family of force fields.

Exotic Species

So, you want to simulate a protein/nucleic acid system, but it binds various exotic metal ions (ruthenium?), or there is an iron-sulfur cluster essential for its functionality, or similar. But, (unfortunately?) there aren’t parameters available for these in the force field you want to use. What should you do? You shoot an e-mail to the GROMACS users emailing list, and get referred to the FAQs.

If you really insist on simulating these in molecular dynamics, you’ll need to obtain parameters for them, either from the literature, or by doing your own parametrization. But before doing so, it’s probably important to stop and think, as sometimes there is a reason there may not already be parameters for such atoms/clusters. In particular, here are a couple of basic questions you can ask yourself to see whether it’s reasonable to develop/obtain standard parameters for these and use them in molecular dynamics:

  • Are quantum effects (i.e. charge transfer) likely to be important? (i.e., if you have a divalent metal ion in an enzyme active site and are interested in studying enzyme functionality, this is probably a huge issue).
  • Are standard force field parametrization techniques used for my force field of choice likely to fail for an atom/cluster of this type? (i.e. because Hartree-Fock 6-31G* can’t adequately describe transition metals, for example)

If the answer to either of these questions is “Yes”, you may want to consider doing your simulations with something other than classical molecular dynamics.

Even if the answer to both of these is “No”, you probably want to consult with someone who is an expert on the compounds you’re interested in, before attempting your own parametrization. Further, you probably want to try parametrizing something more straightforward before you embark on one of these.

Potential of Mean Force

The potential of mean force (PMF) is defined as the potential that gives an average force over all the configurations of a given system. There are several ways to calculate the PMF in GROMACS, probably the most common of which is to make use of the pull code. The steps for obtaining a PMF using umbrella sampling, which allows for sampling of statistically-improbable states, are:

  • Generate a series of configurations along a reaction coordinate (from a steered MD simulation, a normal MD simulation, or from some arbitrarily-created configurations)
  • Use umbrella sampling to restrain these configurations within sampling windows.
  • Use gmx wham to make use of the WHAM algorithm to reconstruct a PMF curve.

A more detailed tutorial is linked here for umbrella sampling.

Single-Point Energy

Computing the energy of a single configuration is an operation that is sometimes useful. The best way to do this with GROMACS is with the mdrun -rerun mechanism, which applies the model physics in the tpr to the configuration in the trajectory or coordinate file supplied to mdrun.

mdrun -s input.tpr -rerun configuration.pdb

Note that the configuration supplied must match the topology you used when generating the tpr file with grompp. The configuration you supplied to grompp is irrelevant, except perhaps for atom names. You can also use this feature with energy groups (see the Reference manual), or with a trajectory of multiple configurations (and in this case, by default mdrun will do neighbour searching for each configuration, because it can make no assumptions about the inputs being similar).

A zero-step energy minimization does a step before reporting the energy, and a zero-step MD run has (avoidable) complications related to catering to possible restarts in the presence of constraints, so neither of those procedures are recommended.

Carbon Nanotube

Robert Johnson’s Tips

Taken from Robert Johnson’s posts on the gmx-users mailing list.

  • Be absolutely sure that the “terminal” carbon atoms are sharing a bond in the topology file.
  • Use periodic_molecules = yes in your mdp file for input in gmx grompp.
  • Even if the topology is correct, crumpling may occur if you place the nanotube in a box of wrong dimension, so use VMD to visualize the nanotube and its periodic images and make sure that the space between images is correct. If the spacing is too small or too big, there will be a large amount of stress induced in the tube which will lead to crumpling or stretching.
  • Don’t apply pressure coupling along the axis of the nanotube. In fact, for debugging purposes, it might be better to turn off pressure coupling altogether until you figure out if anything is going wrong, and if so, what.
  • When using x2top with a specific force field, things are assumed about the connectivity of the molecule. The terminal carbon atoms of your nanotube will only be bonded to, at most, 2 other carbons, if periodic, or one if non-periodic and capped with hydrogens.
  • You can generate an “infinite” nanotube with the -pbc option to x2top. Here, x2top will recognize that the terminal C atoms actually share a chemical bond. Thus, when you use grompp you won’t get an error about a single bonded C.

Andrea Minoia’s tutorial

Modeling Carbon Nanotubes with GROMACS (also archived as contains everything to set up simple simulations of a CNT using OPLS-AA parameters. Structures of simple CNTs can be easily generated e.g. by buildCstruct (Python script that also adds terminal hydrogens) or TubeGen Online (just copy and paste the PDB output into a file and name it cnt.pdb).

To make it work with modern GROMACS you’ll probably want to do the following:

  • make a directory cnt_oplsaa.ff
  • In this directory, create the following files, using the data from the tutorial page:
    • forcefield.itp from the file in section itp
    • atomnames2types.n2t from the file in section n2t
    • aminoacids.rtp from the file in section rtp
  • generate a topology with the custom forcefield (the cnt_oplsaa.ff directory must be in the same directory as where the gmx x2top command is run or it must be found on the GMXLIB path), -noparam instructs gmx x2top to not use bond/angle/dihedral force constants from the command line (-kb, -ka, -kd) but rely on the force field files; however, this necessitates the next step (fixing the dihedral functions)
gmx x2top -f cnt.gro -o -ff cnt_oplsaa -name CNT -noparam

The function type for the dihedrals is set to ‘1’ by gmx x2top but the force field file specifies type ‘3’. Therefore, replace func type ‘1’ with ‘3’ in the [ dihedrals ] section of the topology file. A quick way is to use sed (but you might have to adapt this to your operating system; also manually look at the top file and check that you only changed the dihedral func types):

sed -i~ '/\[ dihedrals \]/,/\[ system \]/s/1 *$/3/'

Once you have the topology you can set up your system. For instance, a simple in-vacuo simulation (using your favourite parameters in em.mdp and md.mdp):

Put into a slightly bigger box:

gmx editconf -f cnt.gro -o boxed.gro -bt dodecahedron -d 1

Energy minimise in vacuuo:

gmx grompp -f em.mdp -c boxed.gro -p -o em.tpr
gmx mdrun -v -deffnm em

MD in vacuuo:

gmx grompp -f md.mdp -c em.gro -p -o md.tpr
gmx mdrun -v -deffnm md

Look at trajectory:

gmx trjconv -f md.xtc -s md.tpr -o md_centered.xtc -pbc mol -center
gmx trjconv -s md.tpr -f md_centered.xtc -o md_fit.xtc -fit rot+trans
vmd em.gro md_fit.xtc