High-throughput design of symmetrical dimeric SARS-CoV-2 main protease: structural and physical insights into hotspots for adaptation and therapeutics.

Dimerization of SARS-CoV-2 main protease (M pro ) is a prerequisite for its processing activity. With >2000 mutations already reported in M pro , SARS-CoV-2 may accumulate mutations in the M pro dimeric interface to stabilize it further. We employed high-throughput protein design strategies to design the symmetrical dimeric interface of M pro (300 000 designs) to identify mutational hotspots that render the M pro more stable. We found that ∼22% of designed mutations that yield stable M pro dimers already exist in SARS-CoV-2 genomes and are currently circulating. Our multi-parametric analyses highlight potential M pro mutations that SARS-CoV-2 may develop, providing a foundation for assessing viral adaptation and mutational surveillance.