Dynamical Nonequilibrium Molecular Dynamics Simulations Identify Allosteric Sites and Positions Associated with Drug Resistance in the SARS-CoV-2 Main Protease.

The SARS-CoV-2 main protease (M) plays an essential role in the coronavirus lifecycle by catalyzing hydrolysis of the viral polyproteins at specific sites. M is the target of drugs, such as nirmatrelvir, though resistant mutants have emerged that threaten drug efficacy. Despite its importance, questions remain on the mechanism of how M binds its substrates. Here, we apply dynamical nonequilibrium molecular dynamics (D-NEMD) simulations to evaluate structural and dynamical responses of M to the presence and absence of a substrate. The results highlight communication between the M dimer subunits and identify networks, including some far from the active site, that link the active site with a known allosteric inhibition site, or which are associated with nirmatrelvir resistance. They imply that some mutations enable resistance by altering the allosteric behavior of M. More generally, the results show the utility of the D-NEMD technique for identifying functionally relevant allosteric sites and networks including those relevant to resistance.