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A Fluorogenic Peptide Cleavage Assay to Screen for Proteolytic Activity: Applications for coronavirus spike protein activation
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Structure of coronavirus main proteinase reveals combination of a chymotrypsin fold with an extra alpha-helical

Kanchan Anand1, Gottfried J Palm, Jeroen R Mesters

  • 1Department of Structural Biology and Crystallography, Institute of Molecular Biotechnology, D-07745 Jena , Germany.

The EMBO Journal
|July 3, 2002
PubMed

Insights

The crystal structure of the transmissible gastroenteritis virus main proteinase (Mpro) reveals a unique chymotrypsin-like fold. This structure explains how Mpro self-processes, offering insights into coronavirus replication mechanisms.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Virology

Background:

  • The viral main proteinase (Mpro) is crucial for processing coronavirus polyproteins.
  • Mpro exhibits low sequence similarity to other known proteinases.

Purpose of the Study:

  • To determine the crystal structure of the transmissible gastroenteritis virus Mpro.
  • To elucidate the mechanism of Mpro self-processing and substrate binding.

Main Methods:

  • X-ray crystallography to determine the 3D structure of Mpro.
  • Molecular modeling and mutagenesis to investigate substrate interactions.

Main Results:

  • The 33.1 kDa Mpro structure was refined to 1.96 A resolution.
  • Three dimers were observed in the asymmetric unit, suggesting trans-cleavage activity.
  • A novel alpha-helical domain and a substrate-binding loop were identified.
  • Specific subsites (S1 and S2) were characterized for binding P1 glutamine and P2 leucine residues.

Conclusions:

  • The study reveals an unprecedented variation of the chymotrypsin fold in RNA viruses.
  • Interactions involving the N-terminus and alpha-helical domain stabilize the active conformation for trans-cleavage.
  • The findings provide a structural basis for understanding coronavirus polyprotein processing.

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