An extended conformation of SARS-CoV-2 main protease reveals allosteric targets

The coronavirus main protease (M-pro) is required for viral replication and has enzymatical activity as a homodimer. Thus, targeting its dimerization is an effective strategy for developing allosteric inhibitors to suppress mutation escape. In this study, we obtained the extended conformation of the native monomer of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) M-pro by trapping it with nanobodies, and found that the catalytic domain and the helix domain dissociate, revealing allosteric targets. We also found another state, a compact conformation, similar to the dimeric form. Our data support that the M-pro may be in equilibrium among the monomeric extended conformation as the precursor of all other states, the compact conformation as the intermediate state, and the dimeric conformation as the active state. We designed an innovative Nanoluc Binary Technology (NanoBiT)-based high-throughput allosteric inhibitor assay based on the rearranged conformation. In addition, we identified a set of allosteric inhibitory nanobodies against M-pro, one of which is also a competitive inhibitor of M-pro. Our results provide insight into the maturation of the coronavirus M-pro and a way to develop anticoronaviral drugs through targeting the folding process to inhibit the autocleavage of the main protease.

Automated summary

This study reveals the structure and function of the coronavirus M-proteinase, showing that it forms an active state through a folding process, providing a new approach for developing antiviral drugs against coronaviruses.