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Unusual architecture of the p7 channel from hepatitis C virus

Nature volume 498, pages 521–525 (2013)Cite this article

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Abstract

The hepatitis C virus (HCV) has developed a small membrane protein, p7, which remarkably can self-assemble into a large channel complex that selectively conducts cations1,2,3,4. We wanted to examine the structural solution that the viroporin adopts in order to achieve selective cation conduction, because p7 has no homology with any of the known prokaryotic or eukaryotic channel proteins. The activity of p7 can be inhibited by amantadine and rimantadine2,5, which are potent blockers of the influenza M2 channel6 and licensed drugs against influenza infections7. The adamantane derivatives have been used in HCV clinical trials8, but large variation in drug efficacy among the various HCV genotypes has been difficult to explain without detailed molecular structures. Here we determine the structures of this HCV viroporin as well as its drug-binding site using the latest nuclear magnetic resonance (NMR) technologies. The structure exhibits an unusual mode of hexameric assembly, where the individual p7 monomers, i, not only interact with their immediate neighbours, but also reach farther to associate with the i+2 and i+3 monomers, forming a sophisticated, funnel-like architecture. The structure also points to a mechanism of cation selection: an asparagine/histidine ring that constricts the narrow end of the funnel serves as a broad cation selectivity filter, whereas an arginine/lysine ring that defines the wide end of the funnel may selectively allow cation diffusion into the channel. Our functional investigation using whole-cell channel recording shows that these residues are critical for channel activity. NMR measurements of the channel–drug complex revealed six equivalent hydrophobic pockets between the peripheral and pore-forming helices to which amantadine or rimantadine binds, and compound binding specifically to this position may allosterically inhibit cation conduction by preventing the channel from opening. Our data provide a molecular explanation for p7-mediated cation conductance and its inhibition by adamantane derivatives.

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Figure 1: NMR structure of the p7(5a) hexamer and its comparison to the EM map.
Figure 2: The pore properties of the p7(5a) channel.
Figure 3: NMR characterization of the amantadine binding site.
Figure 4: A model for amantadine or rimantadine inhibition of the p7 channel.

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Accession codes

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Protein Data Bank

Data deposits

The structure is deposited in the Protein Data Bank under the accession number 2M6X.

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Acknowledgements

We thank R. Sounier for helping with making specific methyl-labelled protein, S. Brueschweiler for helping with ITC measurements, G. Bellot, J. Min and W. Shih for providing DNA nanotube liquid crystal, and K. Oxenoid for discussion. This work was supported by the National Key Project of 973 (2013CB530504) and National Science and Technology Major Project (2012ZX10002-007-003) (to B.S.) and NIH grant GM094608 (to J.J.C.).

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Authors and Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Bo OuYang, Marcelo J. Berardi, Jyoti Dev & James J. Chou

  2. State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China,

    Bo OuYang & James J. Chou

  3. National Center for Protein Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China,

    Bo OuYang & James J. Chou

  4. Molecular Virus Unit, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200025, China,

    Shiqi Xie, Xinhao Zhao, Wenjing Yu & Bing Sun

  5. State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, 200031, China

    Bing Sun

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Contributions

B.O. and J.J.C. conceived the study; B.O. prepared samples; M.J.B. performed EM analysis; J.D. and B.O. performed NMR titration; B.O. and J.J.C. collected and analysed NMR data and determined the structure; S.X., X.Z., W.Y. and B.S. designed and performed functional experiments; J.J.C. wrote the paper and all authors contributed to the editing of the manuscript.

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Correspondence to Bing Sun or James J. Chou.

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The authors declare no competing financial interests.

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OuYang, B., Xie, S., Berardi, M. et al. Unusual architecture of the p7 channel from hepatitis C virus. Nature 498, 521–525 (2013). https://doi.org/10.1038/nature12283

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