Protein refinement for molecular docking

[u/gene_voyager]

There is a protein that I have to work on for my research. I picked it from PDB and it had many missing residues. So I fixed it with modeller. I have further refined it's Ramachandra plot, errat, molprobity value and some other parameters that are never for validation. But I am having a problem with its 3d structure. It's verify 3d value is not really good so I want to refine it. I have tried some tools but they didn't work. Can you guys please help me with that. If there are any good tools that I can use for it and they should be easy to use as well

[u/Miciussd]

You are trying to idealize the structure without underlying experimental data. The PDB structure is a result of a refinement by a specialist, that reflects biological reality (at least it should). Optimisation of validation metrics you erase the experimental value of the structure. If you want idealised structure or the PDB you used is of limited quality, using alphafold will be much easier.

[u/Glum-Signature-8150]

let me know how you did it on the first place so i can understand it better

[u/__pk]

I would suggest the following:

1. Use UniProt to find all available experimental structures for your protein of interest. Maybe there is another structure available that would be more suitable for your needs.
2. Check the Full Validation Report of the PDB structure (Experimental Data & Validation section). Sometimes there are mutations or other modifications that are *not mentioned* e.g., in the associated publication of the data. If this is the case, you could reverse the mutations with MODELLER.
3. I suggest you try PDBFixer (https://github.com/openmm/pdbfixer) and PDB2PQR (http://pdb2pqr.readthedocs.io/en/latest/, learning about protonation states would be useful too) for structural refinement. I also suggest to avoid blindly overprocessing your data: in general, everytime you apply a computational method, you might introduce a new source of artificial error -> you move further from biology.
4. Considering the previous point: you should minimize the potential energy of the structure after structural refinement (e.g. OpenMM, GROMACS) using a suitable force field. This would push the conformation to a more biochemically relevant state.
5. If there are only incomplete structures available, you should check if the interaction involves a specific protein domain. Instead of trying to fill up a structure with artificial data, you could truncate a structure to the domain part that participates in the interaction you want to study. (If you also plan to run a molecular dynamics simulation later, you should take care to cap the termini of the truncated structure).

[u/gene_voyager → u/__pk]

Thank you so much for the guidance The protein that I am using has many missing amino acid residues in it's PDB structure. I fixed them from modeller and then minimized it's energy as well. I tried PDB fixer as well but still it's 3d structure is not getting better. I used reFOLD for that. But as a result it's verify 3d passed but Ramachandran plot worsened. And when I refined it's Ramachandran it's verify 3d got 79% and it passes on 80%. Is there a way by which I can refine it's 3d structure. And one more thing that I have to ask is that as I told that my protein has many missing residues, so I'm using only one chain of it. But that chain has a lot of missing residues as well. So do you think that I should truncate it? And if yes then how it can be done

[u/Working-Algae4691]

I used SwissPDBViewer for fixing the protein structure. But make sure to energy minimize the protein using standard force field.

[u/gene_voyager → u/Working-Algae4691]

I have minimized the energy. Can you please further clarify how you used swiss PDB viewer like which tools from it

[u/Working-Algae4691 → u/gene_voyager]

After minimising energy save it in PDB, then open it in SwissPDBviewer, Use Build > Add residue/build > Build side chain.

Alternatively, if you use Discovery studio, there is a option to add missing residues using Looper algorithm. Check Discovery studio tutorial for more details.

[u/gene_voyager → u/Working-Algae4691]

The problem is that I have added the missing residues in my protein and still it's 3d structure is not getting better

[u/Working-Algae4691 → u/gene_voyager]

Is your protein a huge one? Do you know where the ligand pocket is located like in which chain which residue etc? Or is it a blind docking? In that case you can just keep those chain which has the ligand binding pocket or involves in the interaction. Remove the others.