Multiple protonation states in ligand-free SARS-CoV-2 main protease revealed by large-scale quantum molecular dynamics simulations.

The main protease (Mpro) in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) catalyzes the cleavage of polyproteins for viral replication. Here, large-scale quantum molecular dynamics and metadynamics simulations for ligand-free Mpro were performed, where all the atoms were treated quantum-mechanically, focusing on elucidation of the controversial active-site protonation state. The simulations clarified that the interconverting multiple protonation states exist in unliganded Mpro, and the catalytically relevant ion-pair state is more stable than the neutral state, which is consistent with neutron crystallography. The results highlight the importance of the ion-pair state for repurposing or discovering antiviral drugs that target Mpro.

Asymmetric dynamics of dimeric SARS-CoV-2 and SARS-CoV main proteases in an apo form: Molecular dynamics study on fluctuations of active site, catalytic dyad, and hydration water.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been widely spread around the world. It is necessary to examine the viral proteins that play a notorious role in the invasion of our body. The main protease (3CLpro) facilitates the maturation of the coronavirus. It is thought that the dimerization of 3CLpro leads to its catalytic activity; the detailed mechanism has, however, not been suggested. Furthermore, the structural differences between the predecessor SARS-CoV 3CLpro and SARS-CoV-2 3CLpro have not been fully understood. Here, we show the structural and dynamical differences between the two main proteases, and demonstrate the relationship between the dimerization and the activity via atomistic molecular dynamics simulations. Simulating monomeric and dimeric 3CLpro systems for each protease, we show that (i) global dynamics between the two different proteases are not conserved, (ii) the dimerization stabilizes the catalytic dyad and hydration water molecules behind the dyad, and (iii) the substrate-binding site (active site) and hydration water molecules in each protomer fluctuate asymmetrically. We then speculate the roles of hydration water molecules in their catalytic activity.