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Discovery of an allosteric mechanism for the regulation of HCV NS3 protein function

Nature Chemical Biology volume 8, pages 920–925 (2012)Cite this article

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Abstract

Here we report a highly conserved new binding site located at the interface between the protease and helicase domains of the hepatitis C virus (HCV) NS3 protein. Using a chemical lead, identified by fragment screening and structure-guided design, we demonstrate that this site has a regulatory function on the protease activity via an allosteric mechanism. We propose that compounds binding at this allosteric site inhibit the function of the NS3 protein by stabilizing an inactive conformation and thus represent a new class of direct-acting antiviral agents.

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Figure 1: Crystal structure of the full-length NS3–NS4a protein.
Figure 2: Cocrystal structures of fragment hits.
Figure 3: Protein-ligand co-crystal structures of compounds 3–5, which were generated using structure-guided optimization.
Figure 4: Proposed mode of action of allosteric inhibitors.

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Acknowledgements

We acknowledge the contributions of M. Vinkovic, H. Angove, G.E. Besong, F. Holding, E. Chiarparin and M.L.Verdonk. We thank C. Cartwright for producing figures. X-ray data were collected at the European Synchrotron Radiation Facility and at the Diamond Lightsource. Sedimentation velocity experiments were performed by C. May and T. Laue at the Center to Advance Molecular Interaction Science at the University of New Hampshire. This research was supported by grant 087738 from the Wellcome Trust.

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Authors and Affiliations

  1. Astex Pharmaceuticals, Cambridge Science Park, Cambridge, UK

    Susanne M Saalau-Bethell, Andrew J Woodhead, Gianni Chessari, Maria G Carr, Joseph Coyle, Brent Graham, Steven D Hiscock, Christopher W Murray, Puja Pathuri, Sharna J Rich, Caroline J Richardson, Pamela A Williams & Harren Jhoti

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  1. Susanne M Saalau-Bethell
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Contributions

S.M.S.-B. performed construct design and expression, protein purification, characterization and crystallization. G.C., A.J.W., M.G.C., S.D.H. and C.W.M. modeled and designed the compounds. A.J.W., M.G.C. and S.D.H. synthesized and characterized the compounds. J.C. performed ITC experiments. B.G. executed the western blot analysis and subgenomic replicon experiments and generated and characterized the resistance mutations. P.P. and P.A.W. crystallized the protein, collected the X-ray data and solved the crystal structures. S.J.R. and C.J.R. performed bioassay experiments. S.M.S.-B. and H.J. conceived the project. S.M.S.-B., A.J.W. and H.J. wrote the manuscript and managed the project. S.M.S-B., J.C., C.W.M. and H.J. conceived the model and designed the experiments.

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Correspondence to Harren Jhoti.

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All authors are employees of Astex Pharmaceuticals and own shares in the company.

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Saalau-Bethell, S., Woodhead, A., Chessari, G. et al. Discovery of an allosteric mechanism for the regulation of HCV NS3 protein function. Nat Chem Biol 8, 920–925 (2012). https://doi.org/10.1038/nchembio.1081

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