Letter | Published:

HIV-1 uses dynamic capsid pores to import nucleotides and fuel encapsidated DNA synthesis

Nature volume 536, pages 349353 (18 August 2016) | Download Citation

Abstract

During the early stages of infection, the HIV-1 capsid protects viral components from cytosolic sensors and nucleases such as cGAS and TREX, respectively, while allowing access to nucleotides for efficient reverse transcription1. Here we show that each capsid hexamer has a size-selective pore bound by a ring of six arginine residues and a ‘molecular iris’ formed by the amino-terminal β-hairpin. The arginine ring creates a strongly positively charged channel that recruits the four nucleotides with on-rates that approach diffusion limits. Progressive removal of pore arginines results in a dose-dependent and concomitant decrease in nucleotide affinity, reverse transcription and infectivity. This positively charged channel is universally conserved in lentiviral capsids despite the fact that it is strongly destabilizing without nucleotides to counteract charge repulsion. We also describe a channel inhibitor, hexacarboxybenzene, which competes for nucleotide binding and efficiently blocks encapsidated reverse transcription, demonstrating the tractability of the pore as a novel drug target.

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Accessions

Data deposits

Atomic coordinates and structure factor files have been deposited in the Protein Data Bank under accession numbers 5HGK, 5HGL, 5HGM, 5HGN, 5HGO, and 5JPA.

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Acknowledgements

This work was funded by the Medical Research Council (UK; U105181010), the European Research Council (281627 -IAI), the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007-2013), ERC grant agreement number 339223 and the National Institute for Health Research University College London Hospitals Biomedical Research Centre. G.J.T. was supported by a Wellcome Trust Senior Biomedical Research Fellowship, D.A.J. by an NHMRC Early Career Fellowship (CJ Martin) (GNT1036521) and AJP by a Research Fellowship from Emmanuel College, Cambridge. We thank L. McKeane for her help designing figures.

Author information

Author notes

    • Amanda J. Price

    Present address: Astex Pharmaceuticals, 436 Cambridge Science Park, Milton Road, Cambridge, CB4 0QA, UK.

Affiliations

  1. MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge CB2 0QH, UK

    • David A. Jacques
    • , William A. McEwan
    • , Amanda J. Price
    •  & Leo C. James
  2. Infection and Immunity, University College London, Cruciform Building 3.3, 90 Gower Street, London WC1E 6BT, UK

    • Laura Hilditch
    •  & Greg J. Towers

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Contributions

D.A.J. performed the majority of the protein production, crystallization experiments and analysis; the fluorescence anisotropy binding experiments; differential scanning fluorimetry; chimaeric virus production and associated infectivity and RT measurements; and TRIM5 abrogation assay. W.A.M. performed the HIV core preparation and endogenous RT experiments. L.H. performed R18G and H12Y infectivity and RT characterizations. A.J.P. crystallized and collected diffraction data from CAhexamer in the open state. L.C.J. performed the stopped-flow kinetics experiments. G.J.T. and L.C.J. supervised the project. The paper was primarily written by D.A.J. and L.C.J. All authors discussed the results and implications and commented on the manuscript at all stages.

Corresponding authors

Correspondence to Greg J. Towers or Leo C. James.

Reviewer Information Nature thanks H. Bayley, P. Cherepanov, M. Yeager and T. Yeates for their contribution to the peer review of this work.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Data

    The file contains the gel source data for Extended Data Figure 5a.

Videos

  1. 1.

    Structural morph between the closed and open states of CAHexamer

    On the left the protein is represented in cartoon format, coloured according to secondary structure. The sidechain of L6 is shown as sticks to emphasise that it is this residue that results in pore closure. On the right P1, H12, T48, Q50, and D51 are represented as sticks to show that the movement of the β-hairpin is driven by the formation of a salt-bridge between H12 and D51. Distances shown are in Ångstroms.

  2. 2.

    Pore opening exposes R18

    Surface representation of the morph depicted in Supplementary Video 1. The β-hairpin and R18 are coloured yellow and blue, respectively.

About this article

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DOI

https://doi.org/10.1038/nature19098

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