Computational Mechanistic Insights into Reversible and Irreversible Covalent Inhibitors Targeting SARS-CoV-2 Mpro

The treatment of SARS-CoV-2 can be accomplished by an effective suppression of its 3CL protease (3CLpro), also known as the main protease (Mpro) and nonstructural protein 5 (nsp5). Covalent inhibitors can irreversibly and selectively disable the protease, particularly when they are highly exothermic. Herein we delve into the distinct kinetic behaviors exhibited by two covalently linked SARS-CoV-2 inhibitors. One of these inhibitors features a nitrile reactive group, while the other has this group replaced by an alkyne group, a less reactive electrophile. Our investigations involve the assessment of pertinent free energy surfaces through the utilization of both ab initio and empirical valence bond (EVB) simulations. The calculated free energy profiles show that substituting the nitrile group with alkyne significantly increases the overall reaction exothermicity. This leads to an efficient inhibition, even though the reaction of the nitrile group has a substantially lower barrier than the alkyne group. We examine the time-dependence of IC50 inhibition by applying a novel kinetic simulation approach, which is particularly important in studies of covalent inhibitors with a very exothermic bonding step. Our computational approach reproduces the observed binding kinetics and appears to provide a powerful tool for studies of covalent inhibitors.