Task 8: Molecular Dynamics Simulations

Task description

A detailed task description can be found here.

Intro and selection of two mutants

The description given here was applied for our wild type (1J8U). However, the steps done for our two mutants R408W and P281L are the same.

We selected R408 we because it is the most abundant mutation of PAH which is associated with phenylketonuria. It has a total frequency of 6.67% (see [1]).

The second mutation, P281L, was selected because it is the closed mutation to the binding site (HIS 285) and we hope that we are able to see some interesting things here which give us an explanation why this mutation is disease causing.

Preperation

Before we can start to generate all necessary files for our MD simulation we have to prepare our structure first. We employed the following steps:

1. Extracting the crystal water below 15 Å

repairPDB 1J8U.pdb -ssw 15 > water_below_15.out

We received the following output:

HETATM 2559  O   HOH A1008       2.996   9.738  16.094  1.00 14.91           O  
HETATM 2561  O   HOH A1010      -3.667  12.788   8.538  1.00 14.80           O  
HETATM 2572  O   HOH A1021       3.557  27.911  10.913  1.00 14.69           O  
HETATM 2879  O   HOH A1328      -5.335  24.271  14.490  1.00 14.88           O  
TER

2. Extract only the protein

repairPDB 1J8U.pdb -nosol > 1J8U_nosol.pdb

3. Extract the amino acid sequence and turn it into lower case letter to give it as a input for SCWRL

repairPDB 1J8U.pdb -seq > 1J8U_seq.txt

vim 1J8U_seq.txt
:%s/.*/\L&/g

4. Use SCWRL to complete the side chains of all amino acids

/apps/scwrl4/Scwrl4 -i 1J8U_nosol.pdb -s 1J8U_seq.txt -o 1J8U_nosol_after_SCWRL.pdb | tee scwrl_1J8U_nosol_after_scwrl.out

5. Remove the hydrogen atoms from the SCWRL output

repairPDB 1J8U_nosol_after_SCWRL.pdb -noh > 1J8U_nosol_after_SCWRL_no_h.pdb

6. concatenate the protein and the crystal water into one file

We just added the output of step 1 to the end of the PDB file 1J8U_nosol_after_SCWRL_no_h.pdb and named this file 1J8U_nosol_after_SCWRL_no_h_merged_crystal_water.pdb

Create important GROMACS files

1. We have to create a TOP (topology) and a PAR (parameter) file with the following command:

pdb2gmx -f 1J8U_nosol_after_SCWRL_no_h_merged_crystal_water.pdb -o 1J8U_nosol_after_SCWRL_no_h_merged_crystal_water.gro -p 1J8U_nosol_after_SCWRL_no_h_merged_crystal_water.top -water tip3p -ff amber03 -vsite hydrogens | tee pdb2gmx.out