2. Generate a solvated protein system

2.1. Generate a topology

Using the modified PDB file (chain A of 4AKE with crystal waters removed), generate a topology file for the CHARMM27 force field together with the TIP3P water model using the gmx pdb2gmx tool:

cd top
gmx pdb2gmx -f ../coord/4ake_a.pdb -o protein.pdb -p 4ake.top -i protein_posre.itp -water tip3p -ff charmm27

Note

Total charge -4.000e (in the next step we will add ions to neutralize the system; we need a net-neutral system to properly handle electrostatics)

2.2. Solvate the protein

2.2.1. Adding water

Create a simulation box with gmx editconf and add solvent with gmx solvate:

cd ../solvation
gmx editconf -f ../top/protein.pdb -o boxed.pdb -c -d 0.5 -bt dodecahedron
gmx solvate -cp boxed.pdb -cs spc216 -p ../top/4ake.top -o solvated.pdb

Attention

In order to reduce the system size and make the simulations run faster we are choosing a very tight box (minimum protein-edge distance 0.5 nm, -d 0.5); for simulations you want to publish this number should be 1.2…1.5 nm so that the electrostatic interactions between copies of the protein across periodic boundaries are sufficiently screened.

gmx solvate updates the number of solvent molecules (“SOL”) in the topology file (check the [ system ] section in top/system.top) [1].

2.2.2. Adding ions

Ions can be added with the gmx genion program in Gromacs.

First, we need a basic TPR file (an empty file is sufficient, just ignore the warnings that gmx grompp spits out by setting -maxwarn 10), then run gmx genion (which has convenient options to neutralize the system and set the concentration (check the help!); gmx genion also updates the topology’s [ system ] section if the top file is provided [1]; it reduces the “SOL” molecules by the number of removed molecules and adds the ions, e.g. “NA” and “CL”).

touch ions.mdp
gmx grompp -f ions.mdp -p ../top/4ake.top -c solvated.pdb -o ions.tpr
printf "SOL" | gmx genion -s ions.tpr -p ../top/4ake.top -pname NA -nname CL -neutral -conc 0.15 -o ionized.pdb

The final output is solvation/ionized.pdb. Check visually in VMD (but note that the dodecahedral box is not represented properly). [2].

Footnotes

[1](1, 2) The automatic modification of the top file by gmx solvate and gmx genion can become a problem if you try to run these commands multiple times and you get error messages later (typically from gmx grompp) that the number of molecules in structure file and the topology file do not agree. In this case you might have to manually delete or adjust the corresponding lines in system.top file.
[2]For notes on how to visualize MD systems with VMD see the notes on Trajectory visualization.