Amber Workshop - tutorial 2 - Section 1: Create the necessary topology and coordinate files

(Note: These tutorials are meant to provide illustrative examples of how to use the AMBER software suite to carry out simulations that can be run on a simple workstation in a reasonable period of time. They do not necessarily provide the optimal choice of parameters or methods for the particular application area.)
Copyright Ross Walker 2006

TUTORIAL 2 - SECTION 1

Using dynamics simulations to estimate binding energetics

By Ross Walker

Stage 1 - Create the necessary topology and coordinate files

As in the DNA tutorial the first stage is to create topology (prmtop) and coordinate (inpcrd) files for the system we wish to simulate. We will again use Xleap to do this. The 1jmq pdb is actually an NMR ensemble of structures. For this tutorial we will be using only the first structure from this pdb file. For this you would normally need to open the pdb file in an editor and strip out everything apart from the first structure. Fortunately to save time I have done this already for you. I have also removed all of the hydrogen atoms from this pdb file since the naming format used here does not conform IUPAC nomenclature. As such a lot of the hydrogen atoms would be seen by xleap to be "extra" atoms for this residue. To solve this I have simply deleted the hydrogens from the file. Xleap will automatically add them back for us at "standard" positions to satisfy the valence requirements. One important point, however, is that having the protons present allows one to check the protonation state of any protonatable residues. By default xleap uses the most common protonation states for these residues. For example for histidine a residue name HIS will by default have epsilon protonation. Thus it is best to go through your protein and ensure that you select the correct protonation state for these residues. Be particularly wary of cysteines, histidines and glutamates. In this case histidine residue 32 had both delta and epsilon hydrogens in the pdb file and so I renamed this residue HIP (for doubly protonated histidine). The rest, in the interests of speed, I have left at the default. If this was a "production" simulation, however, you should go through and carefully check the protonation state of your residues matches what you expect. I also ensured that there was a TER card between the protein and the ligand. Otherwise xleap will assume that these are part of the same chain.

The pdb file you will need is here:

complex.pdb

If you want you can quickly visualise this structure in vmd. We will also require structures for just the peptide and protein. Again you can edit the pdb file to create new pdb files that contain just the residues from the peptide (Residues 51 to 60 - labelled chain P) and the protein (residues 5 to 50 labelled chain A). To save time I have done this for you already:

peptide.pdb protein.pdb

Now we should load xleap in order to create the topology files we require. Fortunately this protein contains no non-standard residues and so this procedure should be fairly painless. As before we will use the FF99 force field. So we start xleap:

xleap -s -f $AMBERHOME/dat/leap/cmd/leaprc.ff99

Now we will load the three pdb files into three separate units:

COMPLEX = loadpdb complex.pdb
PEPTIDE = loadpdb peptide.pdb
PROTEIN = loadpdb protein.pdb

You should check that the number of hydrogens you expected to be added were actually added. For the moment just assume it is correct.

If you want to look at these structures you can always edit the relevant unit in xleap.

Let's quickly check the units to make sure there are no missing parameters:

check COMPLEX
check PEPTIDE
check PROTEIN

Ignore the warnings about close contacts (unless they are very close) this is why we minimise before running MD.

Now, we will be running an implicit solvent simulation here so we don't need to solvate our system. We also don't need to add counterions since our system is neutral. So we can just go ahead and save the topology files.

saveamberparm COMPLEX complex.prmtop complex.inpcrd
saveamberparm PEPTIDE peptide.prmtop peptide.inpcrd
saveamberparm PROTEIN protein.prmtop protein.inpcrd

(complex.prmtop, complex.inpcrd, peptide.prmtop, peptide.inpcrd, protein.prmtop, protein.inpcrd)

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