Dear Nima Razzaghi-Asl

What do you mean with "enough separated" COM distances in summary_distances file? What are you sampling? Is it a problem when you generate the starting positions of the sampling windows? Why do you say they are not distant enough?

There is no 'summary_distances' file generated by GROMACS. Are you referring to the following tutorial?

http://www.mdtutorials.com/gmx/umbrella/index.html

Hi Nicola

No, I am not referring to the mentioned tutorial. To be more informative, I am modeling a ligand enzyme system. I mean that when i want to generate different frames (summary_distances.dat) there is not enough distance between center of masses of two pulling groups (ligand and protein).

Dear Nima Razzaghi-Asl

Are you generating the file 'summary_distances.dat' with the bash script from that tutorial?

You need to make sure that you are computing the right thing, that is that you are extracting the right info for the right groups. That script was made for that precise tutorial and needs some (minimal) tuning if you want to apply it to something else.

From you question it seems to me that your reaction coordinate is some distance among the protein and the ligand. Have you visualized the trajectory with a visualization tool like VMD? Just to be sure that the ligand and the protein are really drifting apart. If not, than there is something wrong in the simulation, probably in the .mdp file when you define the pull settings. In the other case, i.e., if the protein/ligand are correctly spanning the reaction coordinate in your trajectory, than the problem is very likely in the script you are using to extract the coordinates and that generates te 'summary_distances.dat' file.

Best,

Nicola

Thank you Nicola

But the only things which need to be corrected within the batch file are 1) time of simulation and 2) name of the pulling groups; that I have corrected both of them.

May the reason be related to the "restrain of protein"? should i change the topology file that was generated in the primary MD simulation (MD simulation without umbrella sampling)?

For further consideration, I also attach the topology file below.

Regards & thanks

Dear Nima Razzaghi-Asl

I would look at the trajectory first of all, to see if it is clear what is not working. I guess you are pulling the ligand away from the protein. Is it really happening or not?

The topology looks fine, especially since the MD simulations are running, which wouldn't happen with errors in the topology, at least with respect to the definition of the molecules. It could be that you need to put some restraint, but again I would look at the trajectory as first thing, otherwise we are randomly guessing without knowing if the system is evolving as supposed to or not. Look also at the pullx file, which in this case is likely the distance of the COM of the two pulling groups. Is it increasing with time? If not, then something is not working in the pulling.

Best

Nicola

Dear Nicola

Yes, everything seems to be OK. However for further consideration, i attach pullx.xvg, pullf.xvg and also a summary_distances.dat file that i have talked about before.

Thanks