Difference between revisions of "2024 AMBER tutorial 1 with PDBID 2ITO"
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=Introduction= | =Introduction= | ||
− | + | AMBER is a molecular dynamics program that simulate the behavior of a protein/ligand complex of interest. AMBER considers all atoms and models their interactions over time, continuously updating positions and forces in small time increments. For this tutorial, we will focus on 2ITO. | |
+ | |||
+ | =Getting Started= | ||
+ | |||
+ | Update the directory structure to the following: | ||
+ | |||
=Structure= | =Structure= | ||
+ | |||
+ | Similarly to the DOCK tutorial, we need to obtain the structure of the protein and ligand. For the protein, we want to fill in all loops even if it is not near the binding pocket, since AMBER considers all atoms in its calculations. Follow the DOCK tutorial for 2ITO to do this. | ||
+ | |||
+ | For the protein, do not add H or add charge. Save as 2ITO_protein_only_for_AMBER.pdb and 2ITO_protein_only_for_AMBER.mol2. | ||
+ | For the ligand, add H and add charge. Save as 2ITO_ligand_only_for_AMBER.pdb and 2ITO_ligand_only_for_AMBER.mol2. | ||
=Force Field Parameters= | =Force Field Parameters= | ||
+ | |||
+ | Force field parameters are required to calculation the interactions between all atoms. This needs to be done for the ligand. Create the following slurm script as follows: | ||
+ | |||
+ | vi 2ITO_ffp.slurm | ||
+ | |||
+ | and have the following within the file: | ||
+ | |||
+ | Make sure to change LIG -nc 0 based off of your ligand. It is the charge on the ligand. For 2ITO, this charge is +1. | ||
+ | |||
+ | Once the slurm file has run, the following will show up in your directory: | ||
+ | |||
+ | 2ITO_ligand_antechamber.mol2 | ||
+ | ANTECHAMBER_AC.AC0 | ||
+ | ANTECHAMBER_AM1BCC_PRE.AC | ||
+ | ANTECHAMBER_BOND_TYPE.AC0 | ||
+ | sqm.in | ||
+ | sqm.pdb | ||
+ | ANTECHAMBER_AC.AC | ||
+ | ANTECHAMBER_AM1BCC.AC | ||
+ | ANTECHAMBER_BOND_TYPE.AC | ||
+ | ATOMTYPE.INF | ||
+ | sqm.out | ||
+ | |||
+ | 2ITO_ligand_antechamber.mol2 is the file with the parameters we generated. | ||
+ | |||
+ | In the command line, load amber 16: | ||
+ | |||
+ | module load amber/16 | ||
+ | |||
+ | Run the following command to modify the parameters slightly: | ||
+ | |||
+ | parmchk2 -i 2ITO_ligand_antechamber.mol2 -f mol2 -o 2ITO_ligand.am1bcc.frcmod | ||
+ | |||
+ | |||
+ | You will see 2ITO_ligand.am1bcc.frcmod file in your directory. | ||
=TLeap= | =TLeap= | ||
+ | |||
+ | TLeap is a program with AMBER that will prepare the system for MD simulations. It will generate the gas-phase and solvated systems. | ||
+ | |||
+ | Create the file: | ||
+ | vi 2ITO_tleap.in | ||
+ | |||
+ | and add the following to the file: | ||
+ | |||
+ | Create the following slurm file: | ||
+ | vi 2ITO_tleap.slurm | ||
+ | |||
+ | and add the following to the file: | ||
+ | |||
+ | |||
+ | Run the slurm and you should get the following new files in your directory: | ||
+ | 2ITO.complex.parm7 | ||
+ | 2ITO.gas.complex.rst7 | ||
+ | 2ITO.gas.ligand.rst7 | ||
+ | 2ITO.gas.receptor.rst7 | ||
+ | leap.log | ||
+ | 2ITO.gas.complex.pdb | ||
+ | 2ITO.gas.ligand.parm7 | ||
+ | 2ITO.gas.receptor.parm7 | ||
+ | 2ITO.wet.complex.pdb | ||
+ | 2ITO.wet.complex.prmtop | ||
+ | 2ITO.wet.complex.rst7 | ||
+ | tleap_output.txt | ||
+ | |||
+ | You can look at the files you created by moving the files to your local computer. Move 2ITO.wet.complex.rst7 and 2ITO.wet.complex.prmtop. Open chimera and follow the following commands: | ||
+ | |||
+ | Tools → MD/Ensemble Analysis → MD Movie | ||
+ | |||
+ | Click browse and put in the prmtop file. Add the rst7 trajectory file. You should be able to view your system. | ||
=Equilibration= | =Equilibration= | ||
+ | |||
+ | Look at the 2ITO.gas.complex.pdb file that was just generated. Scroll all the way down to see the residues in the complex. In our example, the protein is residues 1-324 and the ligand is 325. | ||
+ | |||
+ | We need to equilibrate the system before running the MD simulations. Note, for files 1-7, set the restraint mask to the entire system (1-325). For files 8-9, set the restraint mask to cover only the receptor (1-324). | ||
+ | |||
+ | vi 01.min.mdin | ||
+ | vi 02.equil.mdin | ||
+ | vi 03.min.mdin | ||
+ | vi 04.min.mdin | ||
+ | vi 05.min.mdin | ||
+ | vi 06.equil.mdin | ||
+ | vi 07.equil.mdin | ||
+ | vi 08.equil.mdin | ||
+ | vi 09.equil.mdin | ||
=Production= | =Production= | ||
+ | |||
+ | The system is all equilibrated and now we can run production run. | ||
+ | |||
+ | vi 10.prod.mdin | ||
+ | |||
+ | and add the following: | ||
+ | |||
+ | Create the slurm file: | ||
+ | |||
+ | production.slurm | ||
+ | |||
+ | and add the following: | ||
+ | |||
+ | Run the slurm script | ||
=Analysis= | =Analysis= | ||
+ | |||
+ | Now we can analyze the simulation by looking at the RMSD, H-Bonds, and MM/GBSA results. | ||
==RMSD== | ==RMSD== | ||
+ | |||
+ | First we want to strip all the water: | ||
+ | |||
+ | vi cpp_strip_water.in | ||
+ | |||
+ | with the following: | ||
+ | |||
+ | |||
+ | To look at the RMSD of the ligand, we only want to look at the residue that includes the ligand: | ||
+ | |||
+ | vi cpp_rmsd_lig.in | ||
+ | |||
+ | with the following: | ||
+ | |||
+ | |||
+ | Run it with the following command: | ||
+ | |||
+ | cpptraj -i cpp_strip_water.in | ||
+ | |||
+ | |||
+ | Run it with the following command: | ||
+ | cpptraj -p ../003_tleap/2ITO.complex.parm7 -i cpp_rmsd_lig.in | ||
+ | |||
+ | |||
+ | Now we want to look at the RMSD of the receptor. Include the residues that include only the receptor: | ||
+ | |||
+ | vi cpp_rmsd_receptor.in | ||
+ | |||
+ | with the following: | ||
+ | |||
+ | |||
+ | Run it with the following command: | ||
+ | cpptraj -p ../003.tleap/4s0v.complex.parm7 -i cpp_rmsd_receptor.in | ||
+ | |||
+ | If you want to look at the histogram, you can graph it using python. | ||
+ | |||
+ | |||
+ | ==H-Bonds== | ||
+ | |||
+ | To look at the hydrogen bonds that occur in the system during the simulation, make sure to include the receptor and ligand in the mask. Create the program: | ||
+ | |||
+ | vi cpp_hbonds.in | ||
+ | |||
+ | with the following: | ||
+ | |||
+ | Run it with the following command: | ||
+ | cpptraj -p ../003_tleap/2ITO.wet.complex.prmtop -i cpp_hbonds.in | ||
+ | |||
==MM/GBSA== | ==MM/GBSA== | ||
+ | |||
+ | Now we will calcualte the free energy of the system. Create the program: | ||
+ | |||
+ | vi mmgbsa_2ITO.in | ||
+ | |||
+ | with the following: | ||
+ | |||
+ | and run. |
Revision as of 17:53, 5 May 2024
Contents
Introduction
AMBER is a molecular dynamics program that simulate the behavior of a protein/ligand complex of interest. AMBER considers all atoms and models their interactions over time, continuously updating positions and forces in small time increments. For this tutorial, we will focus on 2ITO.
Getting Started
Update the directory structure to the following:
Structure
Similarly to the DOCK tutorial, we need to obtain the structure of the protein and ligand. For the protein, we want to fill in all loops even if it is not near the binding pocket, since AMBER considers all atoms in its calculations. Follow the DOCK tutorial for 2ITO to do this.
For the protein, do not add H or add charge. Save as 2ITO_protein_only_for_AMBER.pdb and 2ITO_protein_only_for_AMBER.mol2. For the ligand, add H and add charge. Save as 2ITO_ligand_only_for_AMBER.pdb and 2ITO_ligand_only_for_AMBER.mol2.
Force Field Parameters
Force field parameters are required to calculation the interactions between all atoms. This needs to be done for the ligand. Create the following slurm script as follows:
vi 2ITO_ffp.slurm
and have the following within the file:
Make sure to change LIG -nc 0 based off of your ligand. It is the charge on the ligand. For 2ITO, this charge is +1.
Once the slurm file has run, the following will show up in your directory:
2ITO_ligand_antechamber.mol2 ANTECHAMBER_AC.AC0 ANTECHAMBER_AM1BCC_PRE.AC ANTECHAMBER_BOND_TYPE.AC0 sqm.in sqm.pdb ANTECHAMBER_AC.AC ANTECHAMBER_AM1BCC.AC ANTECHAMBER_BOND_TYPE.AC ATOMTYPE.INF sqm.out
2ITO_ligand_antechamber.mol2 is the file with the parameters we generated.
In the command line, load amber 16:
module load amber/16
Run the following command to modify the parameters slightly:
parmchk2 -i 2ITO_ligand_antechamber.mol2 -f mol2 -o 2ITO_ligand.am1bcc.frcmod
You will see 2ITO_ligand.am1bcc.frcmod file in your directory.
TLeap
TLeap is a program with AMBER that will prepare the system for MD simulations. It will generate the gas-phase and solvated systems.
Create the file:
vi 2ITO_tleap.in
and add the following to the file:
Create the following slurm file:
vi 2ITO_tleap.slurm
and add the following to the file:
Run the slurm and you should get the following new files in your directory:
2ITO.complex.parm7 2ITO.gas.complex.rst7 2ITO.gas.ligand.rst7 2ITO.gas.receptor.rst7 leap.log 2ITO.gas.complex.pdb 2ITO.gas.ligand.parm7 2ITO.gas.receptor.parm7 2ITO.wet.complex.pdb 2ITO.wet.complex.prmtop 2ITO.wet.complex.rst7 tleap_output.txt
You can look at the files you created by moving the files to your local computer. Move 2ITO.wet.complex.rst7 and 2ITO.wet.complex.prmtop. Open chimera and follow the following commands:
Tools → MD/Ensemble Analysis → MD Movie
Click browse and put in the prmtop file. Add the rst7 trajectory file. You should be able to view your system.
Equilibration
Look at the 2ITO.gas.complex.pdb file that was just generated. Scroll all the way down to see the residues in the complex. In our example, the protein is residues 1-324 and the ligand is 325.
We need to equilibrate the system before running the MD simulations. Note, for files 1-7, set the restraint mask to the entire system (1-325). For files 8-9, set the restraint mask to cover only the receptor (1-324).
vi 01.min.mdin vi 02.equil.mdin vi 03.min.mdin vi 04.min.mdin vi 05.min.mdin vi 06.equil.mdin vi 07.equil.mdin vi 08.equil.mdin vi 09.equil.mdin
Production
The system is all equilibrated and now we can run production run.
vi 10.prod.mdin
and add the following:
Create the slurm file:
production.slurm
and add the following:
Run the slurm script
Analysis
Now we can analyze the simulation by looking at the RMSD, H-Bonds, and MM/GBSA results.
RMSD
First we want to strip all the water:
vi cpp_strip_water.in
with the following:
To look at the RMSD of the ligand, we only want to look at the residue that includes the ligand:
vi cpp_rmsd_lig.in
with the following:
Run it with the following command:
cpptraj -i cpp_strip_water.in
Run it with the following command:
cpptraj -p ../003_tleap/2ITO.complex.parm7 -i cpp_rmsd_lig.in
Now we want to look at the RMSD of the receptor. Include the residues that include only the receptor:
vi cpp_rmsd_receptor.in
with the following:
Run it with the following command:
cpptraj -p ../003.tleap/4s0v.complex.parm7 -i cpp_rmsd_receptor.in
If you want to look at the histogram, you can graph it using python.
H-Bonds
To look at the hydrogen bonds that occur in the system during the simulation, make sure to include the receptor and ligand in the mask. Create the program:
vi cpp_hbonds.in
with the following:
Run it with the following command:
cpptraj -p ../003_tleap/2ITO.wet.complex.prmtop -i cpp_hbonds.in
MM/GBSA
Now we will calcualte the free energy of the system. Create the program:
vi mmgbsa_2ITO.in
with the following:
and run.