Characterization of the non-covalent interaction between the PF-07321332 inhibitor and the SARS-CoV-2 main protease

Abstract

We have studied the non-covalent interaction between PF-07321332 and SARS-CoV2 main protease at the atomic level using a computational approach based on extensive molecular dynamics simulations with explicit solvent. PF-07321332, whose chemical structure has been recently disclosed, is a promising oral antiviral clinical candidate with well-established anti-SARS-CoV-2 activity in vitro. The drug, currently in phase III clinical trials in combination with ritonavir, relies on the electrophilic attack of a nitrile warhead to the catalytic cysteine of the protease. Nonbonded interaction between the inhibitor and the residues of the binding pocket, as well as with water molecules on the protein surface, have been characterized using two different force fields and the two possible protonation states of the main protease catalytic dyad HIS41-CYS145. When the catalytic dyad is in the neutral state, the non-covalent binding is likely to be stronger. Molecular dynamics simulations seems to lend support for an inhibitory mechanism in two steps: a first non-covalent addition with the dyad in neutral form and then the formation of the thiolate-imidazolium ion pair and the ligand relocation for finalising the electrophilic attack.

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

“…The present study clarified that the relevant protonation state for the structural stability of the bound inhibitor differs from the ligands, illustrating the importance of evaluating the binding affinity for the appropriate protonation state. Procacci and co-workers carried out the classical MD simulations of SARS-CoV-2 M pro in the ligand-bound form with PF-07321332 for the IP and neutral states using two different kinds of force fields [25] . The authors found that the active-site protonation state and force fields have implications for the inhibitor-bound structure, highlighting the necessity of the QMD simulation in addition to the classical MD simulation based on empirical force fields.…”

“…Recently, docking and/or classical molecular dynamics (MD) simulations for M pro [18] , [19] , [20] , [21] , [22] , [23] , [24] , [25] , starting from the crystal structures captured from X-ray crystallography, have been widely conducted to search for effective inhibitors by evaluating the affinity for the active site of M pro , mainly consisting of the catalytic dyad, i.e., Cys145 and His41 ( Figure 1 a ). As hydrogen/proton is not resolved in the usual X-ray crystallography due to the spatial resolution, the protonation states should be presumed in these simulations.…”

“…As hydrogen/proton is not resolved in the usual X-ray crystallography due to the spatial resolution, the protonation states should be presumed in these simulations. The neutral state is the protonation state of the catalytic dyad usually employed in these simulations, except for the limited case [22] , [24] , [25] , to the best of our knowledge. The neutral state is consistent with the findings from previous theoretical studies based on the quantum-mechanics/molecular-mechanics (QM/MM) method, in which M pro initiates the catalytic cycle via proton transfer from Cys145 to His41 [26] , [27] , indicating that the ligand-free M pro remains in the neutral form as the resting state before binding the substrate or inhibitors.…”

Oral Nirmatrelvir/Ritonavir Therapy for COVID-19: The Dawn in the Dark?

“…Nirmatrelvir has been observed to form tight conjugates with SARS-CoV-2 M pro [ 17 , 18 , 19 , 20 ]. Analysis of the structure of SARS-CoV-2 M pro in complex with nirmatrelvir shows that, in addition to the S-C covalent bond, nirmatrelvir is further stabilized through a network of hydrogen bonds and hydrophobic interactions, which further strengthens its binding to the active site of SARS-CoV-2 M pro [ 17 ].…”

“…Analysis of the structure of SARS-CoV-2 M pro in complex with nirmatrelvir shows that, in addition to the S-C covalent bond, nirmatrelvir is further stabilized through a network of hydrogen bonds and hydrophobic interactions, which further strengthens its binding to the active site of SARS-CoV-2 M pro [ 17 ]. Molecular dynamics simulations provide further support for the inhibitory mechanism of nirmatrelvir toward SARS-CoV-2 M pro , which occurs in two steps: an initial non-covalent addition with the dyad in a neutral form with the formation of the thiolate-imidazolium ion pair, and ligand relocation for finalizing the electrophilic attack [ 18 ]. Despite the presence of lopinavir or ritonavir, nirmatrelvir still exhibited a high binding ability to the active site of SARS-CoV-2 M pro [ 19 ].…”