An allosteric mechanism for potent inhibition of SARS-CoV-2 main proteinase.

The main proteinase (M) of SARS-CoV-2 plays a critical role in cleaving viral polyproteins into functional proteins required for viral replication and assembly, making it a prime drug target for COVID-19. It is well known that noncompetitive inhibition offers potential therapeutic options for treating COVID-19, which can effectively reduce the likelihood of cross-reactivity with other proteins and increase the selectivity of the drug. Therefore, the discovery of allosteric sites of M has both scientific and practical significance. In this study, we explored the binding characteristics and inhibiting process of M activity by two recently reported allosteric inhibitors, pelitinib and AT7519 which were obtained by the X-ray screening experiments, to probe the allosteric mechanism via molecular dynamic (MD) simulations. We found that pelitinib and AT7519 can stably bind to M far from the active site. The binding affinity is estimated to be -24.37 ± 4.14 and - 26.96 ± 4.05 kcal/mol for pelitinib and AT7519, respectively, which is considerably stable compared with orthosteric drugs. Furthermore, the strong binding caused clear changes in the catalytic site of M, thus decreasing the substrate accessibility. The community network analysis also validated that pelitinib and AT7519 strengthened intra- and inter-domain communication of M dimer, resulting in a rigid M, which could negatively impact substrate binding. In summary, our findings provide the detailed working mechanism for the two experimentally observed allosteric sites of M. These allosteric sites greatly enhance the 'druggability' of M and represent attractive targets for the development of new M inhibitors.