8U4Y

Crystal Structure of SARS-CoV-2 Main Protease (Mpro) L50F Mutant

Summary for 8U4Y

• Entry DOI: 10.2210/pdb8u4y/pdb
• Descriptor: 3C-like proteinase nsp5 (2 entities in total)
• Functional Keywords: protease, sars-cov-2, mpro, apo, covid-19, viral protein, hydrolase
• Biological source: Severe acute respiratory syndrome coronavirus 2
• Total number of polymer chains: 2
• Total formula weight: 67719.12

Authors

Kohaal, N.,Lewandowski, E.M.,Wang, J.,Chen, Y. (deposition date: 2023-09-11, release date: 2024-10-09, Last modification date: 2025-02-12)

Primary citation

Lewandowski, E.M.,Zhang, X.,Tan, H.,Jaskolka-Brown, A.,Kohaal, N.,Frazier, A.,Madsen, J.J.,Jacobs, L.M.C.,Wang, J.,Chen, Y.
Distal protein-protein interactions contribute to nirmatrelvir resistance.
Nat Commun, 16:1266-1266, 2025

PubMed Abstract: SARS-CoV-2 main protease, M, is responsible for processing the viral polyproteins into individual proteins, including the protease itself. M is a key target of anti-COVID-19 therapeutics such as nirmatrelvir (the active component of Paxlovid). Resistance mutants identified clinically and in viral passage assays contain a combination of active site mutations (e.g., E166V, E166A, L167F), which reduce inhibitor binding and enzymatic activity, and non-active site mutations (e.g., P252L, T21I, L50F), which restore the fitness of viral replication. To probe the role of the non-active site mutations in fitness rescue, here we use an M triple mutant (L50F/E166A/L167F) that confers nirmatrelvir drug resistance with a viral fitness level similar to the wild-type. By comparing peptide and full-length M protein as substrates, we demonstrate that the binding of M substrate involves more than residues in the active site. Particularly, L50F and other non-active site mutations can enhance the M dimer-dimer interactions and help place the nsp5-6 substrate at the enzyme catalytic center. The structural and enzymatic activity data of M L50F, L50F/E166A/L167F, and others underscore the importance of considering the whole substrate protein in studying M and substrate interactions, and offers important insights into M function, resistance development, and inhibitor design.

PubMed: 39893201
DOI: 10.1038/s41467-025-56651-x

Experimental method

X-RAY DIFFRACTION (2.21 Å)