Unusual RMSD of ligand-protein

User discussions
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1

GROMACS version: 2024.2
GROMACS modification: No

I have run a protein-ligand simulation for which the backbone RMSD of protein (lsq fit of backbone to backbone of protein) and RMSD of ligand (lsq fit of ligand to backbone) have been calculated.

The ligand RMSD being very high (around 9nm), but doesn’t show any major fluctuations or peak, so probably not a trajectory issue. Also, the ligand seems to be thoroughly bound to the receptor during the 100ns simulation when visualized in VMD.

Below is the plot showing ligand RMSD (in red). and protein backbone RMSD (in black).
Could you please shed some light on why the ligand RMSD is so high?

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Thank you!

2

Hi,

I’m not entirely sure about the wording here:

RMSD of ligand (lsq fit of ligand to backbone) have been calculated.

You probably used gmx rms for this. Did you select “Backbone” first and then “Ligand,” or vice versa?

Best regards,
Marius

3

Yes, backbone first, and then ligand for calculation of ligand RMSD.

4

This could explain your observation. With the options you chose, you are calculating the ligand’s RMSD relative to the protein backbone. Consequently, your ligand curve no longer reflects deviations in the ligand’s internal coordinates, but rather the ligand’s displacement with respect to the protein backbone. Did you perform your analysis on the raw trajectory, or did you first correct for periodic boundary conditions? Which reference structure did you take for the RMSD calculation?

Best,
Marius

5

I corrected for periodic boundary condition with “-pbc mol -center -ur compact” flag of gmx trjconv, finally applied “-fit rot+trans”, checked if the ligand is leaving the binding pocket, but it was consistent within it. Then I calculated RMSD on this fitted trajectory file.

6

Okay, but nevertheless I would guess that the actual calculation of your ligand RMSD is relative to the backbone of the protein. If you select Ligand for both the fitting as well as the RMSD calculation the value should become much smaller (depending on the size and flexibility of your ligand). You can also compare your reference structure (using gmx rms -s ) with individual frames from the fitted trajectory (using gmx rms -f ) to pinpoint which motions could cause this large deviations.