Conformational Flexibility of a Short Loop near the Active Site of the SARS-3CLpro is Essential to Maintain Catalytic Activity

Sci Rep. 2016 Feb 16:6:20918. doi: 10.1038/srep20918.

Abstract

The SARS 3C-like proteinase (SARS-3CLpro), which is the main proteinase of the SARS coronavirus, is essential to the virus life cycle. This enzyme has been shown to be active as a dimer in which only one protomer is active. However, it remains unknown how the dimer structure maintains an active monomer conformation. It has been observed that the Ser139-Leu141 loop forms a short 3(10)-helix that disrupts the catalytic machinery in the inactive monomer structure. We have tried to disrupt this helical conformation by mutating L141 to T in the stable inactive monomer G11A/R298A/Q299A. The resulting tetra-mutant G11A/L141T/R298A/Q299A is indeed enzymatically active as a monomer. Molecular dynamics simulations revealed that the L141T mutation disrupts the 3(10)-helix and helps to stabilize the active conformation. The coil-3(10)-helix conformational transition of the Ser139-Leu141 loop serves as an enzyme activity switch. Our study therefore indicates that the dimer structure can stabilize the active conformation but is not a required structure in the evolution of the active enzyme, which can also arise through simple mutations.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Catalysis
  • Catalytic Domain*
  • Coronavirus 3C Proteases
  • Cysteine Endopeptidases / chemistry*
  • Cysteine Endopeptidases / genetics
  • Cysteine Endopeptidases / metabolism
  • Enzyme Activation
  • Hydrophobic and Hydrophilic Interactions
  • Models, Molecular*
  • Molecular Dynamics Simulation
  • Mutation
  • Protein Conformation*
  • Protein Multimerization
  • Protein Subunits / chemistry
  • Protein Subunits / genetics
  • Protein Subunits / metabolism
  • Viral Proteins / chemistry*
  • Viral Proteins / genetics
  • Viral Proteins / metabolism

Substances

  • Protein Subunits
  • Viral Proteins
  • Cysteine Endopeptidases
  • Coronavirus 3C Proteases