<i>In-silico</i> identification of potential peptide inhibitors to disrupt NLRP3 inflammasome complex formation by blocking NLRP3-ASC pyrin-pyrin interactions

The NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome is a well-known and frequently cited regulator of caspase-1 activation. It plays a significant role in several pathophysiological processes and is a major regulator of the innate immune response. A growing amount of scientific evidences for its aberrant activation in various chronic inflammatory diseases attracts a growing interest in the development of new NLRP3 inhibitors. One of the successful strategies used to identify new inhibitors is peptide inhibitors. Compared to small molecule inhibitors, peptide inhibitors show greater selectivity and less toxicity. In this study, we used an <i>in-silico</i> mutagenesis approach to design new peptide inhibitors from reported peptide inhibitor of NLRP3. The sequence of the peptide inhibitor against NLRP3 was searched from the literature and modeled using the online server PEP-FOLD3. The <i>in-silico</i> alanine scanning mutagenesis of the reference peptide revealed that residues, Y23, R28, E6, I26, R20, L19, Q33, K11, L14, and K13 have positive affinity values and are therefore better candidates for substitution to increase binding affinity. By replacing these residues, the affinity of the newly designed peptide inhibitors for the NLRP3 PYD protein was significantly increased. Further, molecular dynamics simulations and binding energy calculations validated the stability and higher binding affinities of the newly designed peptide inhibitors compared to the wild-type peptide inhibitor. Our research revealed that all the suggested peptide inhibitors have higher binding affinities for the NLRP3 protein as compared to the native wild-type peptide inhibitor and could block NLRP3-ASC pyrin-pyrin interaction.