I want design a peptide for inhibition of a binding site and I found hotspot regions but I don't know on what basis I should start to adding residues to my peptide.
(For imagine my complex please look at 3D structure of 3PQY(PDB file))
You can consider CAMPR3 server. There is a guide for peptide design. The APD3 server and DBAASP should also be of great assistance to guide you.
I hope this helps
It looks like you want to use a synthetic peptide to competitively inhibit binding of H2Db-PA224 to TCR 2-8. The best strategy for blocking TCR binding is to mimic the ligand. Usually the ligand is discontinuous, meaning distant regions of the ligand come together via folding to produce a functional binding interface. Functionally reproducing such a structure with a synthetic peptide is unlikely. But the structure you sent suggests you might be lucky (15-20%) and have a continuous epitope.
At the interface HLA exhibits a beta turn between two alpha helices. If you're lucky, all intermolecular contacts contributed by HLA involve that fairly short contiguous stretch. So you need a peptide that folds like that alpha-turn-alpha structure, and also interacts with TCR via similar non-covalent bonding.
As a first step, you might try simply making a fairly long peptide comprising the AA sequence spanning those two helices including the bend. Likely the helices will fold independently, and the bend angle will be stabilized by binding in the pocket. If that lead compound works in your assay, for your next iteration you can try linking the alpha strands to help define the bend angle.
Take a close look at the crystal structure of the complex, and try to identify residues close enough and properly oriented for non-covalent binding. You can do a pretty good job by eye, but software helps. Software can help predict how stably the synthetic peptide will fold and how well it will bind the TCR.
Before you get started, think about your assay. If you want to identify an inhibitor, you need a sensitive assay that can sensitively detect inhibition. If you haven't already done so, I recommend you clone the HLA and the TCR and optimize a molecular binding assay with serial dilution of HLA (ligand). If you try to use a cell assay, you'll have trouble detecting inhibition, because the cells will report/activate with very low receptor occupancy.
John Carter This is a great guideless, this is my first peptide design experience and my supervisor just told me put complementary residues but I couldn't figure out how, but now I have a clear vision.
Thanks.
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