1. Academic Validation
  2. Bioorthogonal dissection of the replicase assembly of hepatitis C virus

Bioorthogonal dissection of the replicase assembly of hepatitis C virus

  • Cell Chem Biol. 2021 Sep 16;28(9):1366-1378.e4. doi: 10.1016/j.chembiol.2021.03.006.
Yang Zhang 1 Shuiye Chen 1 Zhenghong Yuan 2 Zhigang Yi 3
Affiliations

Affiliations

  • 1 Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
  • 2 Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China. Electronic address: zhyuan@shmu.edu.cn.
  • 3 Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China; Shanghai Public Health Clinical Center, Fudan University, Shanghai 201052, China. Electronic address: zgyi@fudan.edu.cn.
Abstract

Positive-strand RNA viruses such as hepatitis C virus (HCV), flaviviruses, and coronaviruses are medically important. Assembly of replicase on host membranes is a conserved replication strategy and an attractive Antiviral target. The mechanisms of replicase assembly are largely unknown, due to the technical difficulties in purifying the replicase and carrying out structural studies. Here, with an HCV replicase assembly surrogate system, we employed a bioorthogonal system to introduce the photolabile unnatural amino into each residue in the cytosolic regions of NS4B and the amphipathic helix (AH) of NS5A. Photocrosslinking enabled visualization of NS4B oligomerization and NS5A dimerization at pinpointed interacting residues and identifying contacting sites among the replicase components. Characterization of the interacting sites revealed hub elements in replicase assembly by docking replicase components to prompt protein-protein interactions. The results provide information about the molecular architecture of the replicase, advancing understanding of the mechanism of replicase assembly.

Keywords

bioorthogonal strategy; hepatitis C virus; positive-strand RNA virus; replicase assembly; replication complex.

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