Hello everyone,

I have been trying to understand the concept of performing docking for repurposing studies. I've watched a couple of tutorial videos about docking with Pyrx, but at some points, I got confused and I wanted to ask if anybody can help me with that. Probably, the answers to my questions are somewhere on the internet, but I haven't found them yet.

1) Firstly, I am not sure how to find appropriate protein files on the Protein Data Bank (PDB). When I search for a specific protein on the PDB, there are numerous files available, and I don't know which one is suitable for my study. Some proteins have been used in previous studies and they share the same ID number, but what about the other proteins? In tutorials, most of them start with looking for file names, such as 2CNK on the Protein Data Bank.

2) Secondly, my question is about the preparation of the protein for docking. Let's say I have chosen a PDB file to work on, but how am I going to prepare it properly for docking? In some tutorials, people first prepare the protein using other programs like Chimera or AutoDock, and then take the modified PDB to Pyrx. However, in some cases, they directly load their protein on PyRx and modify them there. I'm confused about which approach is more accurate.

3)Thirdly, how to find the active site of the protein or should I use center of mass of the protein as a center of grid box.

4) This question also related with the second one, I read that make macromolecule function in PyRx is remove water, add H and charges. if this is true, making dockprep on Chimera and then loading prepared protein on pyrx then making again macromolecule is wrong thing to do?

Thank you for your answers...