8FIG

Room-temperature X-ray structure of SARS-CoV-2 main protease double mutant E290A/R298A in complex with GC373

Summary for 8FIG

• Entry DOI: 10.2210/pdb8fig/pdb
• Descriptor: 3C-like proteinase nsp5, N~2~-[(benzyloxy)carbonyl]-N-{(2S)-1-hydroxy-3-[(3S)-2-oxopyrrolidin-3-yl]propan-2-yl}-L-leucinamide (3 entities in total)
• Functional Keywords: cysteine protease, homodimer, double mutant, hydrolase
• Biological source: Severe acute respiratory syndrome coronavirus 2 (2019-nCoV, SARS-CoV-2)
• Total number of polymer chains: 1
• Total formula weight: 34086.88

Authors

Kovalevsky, A.,Coates, L.,Kneller, D.W. (deposition date: 2022-12-16, release date: 2023-11-22, Last modification date: 2024-10-16)

Primary citation

Aniana, A.,Nashed, N.T.,Ghirlando, R.,Coates, L.,Kneller, D.W.,Kovalevsky, A.,Louis, J.M.
Insights into the mechanism of SARS-CoV-2 main protease autocatalytic maturation from model precursors.
Commun Biol, 6:1159-1159, 2023

PubMed Abstract: A critical step for SARS-CoV-2 assembly and maturation involves the autoactivation of the main protease (MPro) from precursor polyproteins. Upon expression, a model precursor of MPro mediates its own release at its termini rapidly to yield a mature dimer. A construct with an E290A mutation within MPro exhibits time dependent autoprocessing of the accumulated precursor at the N-terminal nsp4/nsp5 site followed by the C-terminal nsp5/nsp6 cleavage. In contrast, a precursor containing E290A and R298A mutations (MPro) displays cleavage only at the nsp4/nsp5 site to yield an intermediate monomeric product, which is cleaved at the nsp5/nsp6 site only by MPro. MPro and the catalytic domain (MPro) fused to the truncated nsp4 region also show time-dependent conversion in vitro to produce MPro and MPro, respectively. The reactions follow first-order kinetics indicating that the nsp4/nsp5 cleavage occurs via an intramolecular mechanism. These results support a mechanism involving an N-terminal intramolecular cleavage leading to an increase in the dimer population and followed by an intermolecular cleavage at the C-terminus. Thus, targeting the predominantly monomeric MPro precursor for inhibition may lead to the identification of potent drugs for treatment.

PubMed: 37957287
DOI: 10.1038/s42003-023-05469-8

Experimental method

X-RAY DIFFRACTION (1.75 Å)