8V7W

Room-temperature X-ray structure of SARS-CoV-2 main protease catalytic domain C145A precursor, residues nsp4(-6)-1-199-6H

Summary for 8V7W

• Entry DOI: 10.2210/pdb8v7w/pdb
• Related: 8V7T
• Descriptor: Replicase polyprotein 1ab (2 entities in total)
• Functional Keywords: viral protease, catalytic domain, cysteine protease, precursor, hydrolase
• Biological source: Severe acute respiratory syndrome coronavirus 2
• Total number of polymer chains: 2
• Total formula weight: 47379.75

Authors

Kovalevsky, A.,Coates, L. (deposition date: 2023-12-04, release date: 2024-05-29, Last modification date: 2024-06-12)

Primary citation

Kovalevsky, A.,Aniana, A.,Coates, L.,Ghirlando, R.,Nashed, N.T.,Louis, J.M.
Visualizing the Active Site Oxyanion Loop Transition Upon Ensitrelvir Binding and Transient Dimerization of SARS-CoV-2 Main Protease.
J.Mol.Biol., 436:168616-168616, 2024

PubMed Abstract: N-terminal autoprocessing from its polyprotein precursor enables creating the mature-like stable dimer interface of SARS-CoV-2 main protease (MPro), concomitant with the active site oxyanion loop equilibrium transitioning to the active conformation (E*) and onset of catalytic activity. Through mutagenesis of critical interface residues and evaluating noncovalent inhibitor (ensitrelvir, ESV) facilitated dimerization through its binding to MPro, we demonstrate that residues extending from Ser1 through Glu14 are critical for dimerization. Combined mutations G11A, E290A and R298A (MPro™) restrict dimerization even upon binding of ESV to monomeric MPro™ with an inhibitor dissociation constant of 7.4 ± 1.6 µM. Contrasting the covalent inhibitor NMV or GC373 binding to monomeric MPro, ESV binding enabled capturing the transition of the oxyanion loop conformations in the absence of a reactive warhead and independent of dimerization. Characterization of complexes by room-temperature X-ray crystallography reveals ESV bound to the E* state of monomeric MPro as well as an intermediate approaching the inactive state (E). It appears that the E* to E equilibrium shift occurs initially from G138-F140 residues, leading to the unwinding of the loop and formation of the 3-helix. Finally, we describe a transient dimer structure of the MPro precursor held together through interactions of residues A5-G11 with distinct states of the active sites, E and E*, likely representing an intermediate in the autoprocessing pathway.

PubMed: 38762033
DOI: 10.1016/j.jmb.2024.168616

Experimental method

X-RAY DIFFRACTION (1.9 Å)