Multiscale Simulations of the Covalent Inhibition of the SARS-CoV-2 Main Protease: Four Compounds and Three Reaction Mechanisms.

We report the results of computational modeling of the reactions of the SARS-CoV-2 main protease (M Pro ) with four potential covalent inhibitors. Two of them, carmofur and nirmatrelvir, have shown experimentally the ability to inhibit M Pro . Two other compounds, X77A and X77C, were designed computationally in this work. They were derived from the structure of X77, a non-covalent inhibitor forming a tight surface complex with M Pro . We modified the X77 structure by introducing warheads capable of reacting with the catalytic cysteine residue in the M Pro active site. The reaction mechanisms of the four molecules with M Pro were investigated by quantum mechanics/molecular mechanics (QM/MM) simulations. The results show that all four compounds form covalent adducts with the catalytic cysteine Cys 145 of M Pro . From the chemical perspective, the reactions of these four molecules with M Pro follow three distinct mechanisms. The reactions are initiated by a nucleophilic attack of the thiolate group of the deprotonated cysteine residue from the catalytic dyad Cys145-His41 of M Pro . In the case of carmofur and X77A, the covalent binding of the thiolate to the ligand is accompanied by the formation of the fluoro-uracil leaving group. The reaction with X77C follows the nucleophilic aromatic substitution S N Ar mechanism. The reaction of M Pro with nirmatrelvir (which has a reactive nitrile group) leads to the formation of a covalent thioimidate adduct with the thiolate of the Cys145 residue in the enzyme active site. Our results contribute to the ongoing search for efficient inhibitors of the SARS-CoV-2 enzymes.