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


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.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


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.


  1. 1.

    & HIV-1 capsid: the multifaceted key player in HIV-1 infection. Nat. Rev. Microbiol. 13, 471–483 (2015)

  2. 2.

    et al. HIV-1 evades innate immune recognition through specific cofactor recruitment. Nature 503, 402–405 (2013)

  3. 3.

    et al. Host cofactors and pharmacologic ligands share an essential interface in HIV-1 capsid that is lost upon disassembly. PLoS Pathog. 10, e1004459 (2014)

  4. 4.

    et al. HIV-1 DNA Flap formation promotes uncoating of the pre-integration complex at the nuclear pore. EMBO J. 26, 3025–3037 (2007)

  5. 5.

    et al. Crystal structure of human cyclophilin A bound to the amino-terminal domain of HIV-1 capsid. Cell 87, 1285–1294 (1996)

  6. 6.

    et al. Implications for viral capsid assembly from crystal structures of HIV-1 Gag(1-278) and CA(N)(133-278). Biochemistry 45, 11257–11266 (2006)

  7. 7.

    et al. Conformational adaptation of Asian macaque TRIMCyp directs lineage specific antiviral activity. PLoS Pathog. 6, e1001062 (2010)

  8. 8.

    et al. X-ray structures of the hexameric building block of the HIV capsid. Cell 137, 1282–1292 (2009)

  9. 9.

    et al. Structure of the HIV-1 full-length capsid protein in a conformationally trapped unassembled state induced by small-molecule binding. J. Mol. Biol. 406, 371–386 (2011)

  10. 10.

    , & Atomic-level modelling of the HIV capsid. Nature 469, 424–427 (2011)

  11. 11.

    et al. CPSF6 defines a conserved capsid interface that modulates HIV-1 replication. PLoS Pathog. 8, e1002896 (2012)

  12. 12.

    et al. Proteolytic refolding of the HIV-1 capsid protein amino-terminus facilitates viral core assembly. EMBO J. 17, 1555–1568 (1998)

  13. 13.

    , , & Frequent incorporation of ribonucleotides during HIV-1 reverse transcription and their attenuated repair in macrophages. J. Biol. Chem. 287, 14280–14288 (2012)

  14. 14.

    , , & The Moderately Efficient Enzyme: Futile Encounters and Enzyme Floppiness. Biochemistry 54, 4969–4977 (2015)

  15. 15.

    & Rapid, electrostatically assisted association of proteins. Nat. Struct. Biol. 3, 427–431 (1996)

  16. 16.

    , , , & Contribution of unusual arginine-arginine short-range interactions to stabilization and recognition in proteins. J. Protein Chem. 13, 195–215 (1994)

  17. 17.

    , & Unusual arginine formations in protein function and assembly: rings, strings, and stacks. J. Phys. Chem. B 116, 7006–7013 (2012)

  18. 18.

    et al. Structure of B-MLV capsid amino-terminal domain reveals key features of viral tropism, gag assembly and core formation. J. Mol. Biol. 376, 1493–1508 (2008)

  19. 19.

    et al. Extreme genetic fragility of the HIV-1 capsid. PLoS Pathog. 9, e1003461 (2013)

  20. 20.

    , & The human and African green monkey TRIM5alpha genes encode Ref1 and Lv1 retroviral restriction factor activities. Proc. Natl Acad. Sci. USA 101, 10780–10785 (2004)

  21. 21.

    & In vitro uncoating of HIV-1 cores. J. Vis. Exp. 57, 3384 (2011)

  22. 22.

    , & Strand transfer and elongation of HIV-1 reverse transcription is facilitated by cell factors in vitro. PLoS One 5, e13229 (2010)

  23. 23.

    , & Stochastic sensing of nanomolar inositol 1,4,5-trisphosphate with an engineered pore. Chem. Biol. 9, 829–838 (2002)

  24. 24.

    , & Structure and mechanisms of a protein-based organelle in Escherichia coli. Science 327, 81–84 (2010)

  25. 25.

    et al. Selective molecular transport through the protein shell of a bacterial microcompartment organelle. Proc. Natl Acad. Sci. USA 112, 2990–2995 (2015)

  26. 26.

    et al. Protein structures forming the shell of primitive bacterial organelles. Science 309, 936–938 (2005)

  27. 27.

    et al. Active site remodeling switches HIV specificity of antiretroviral TRIMCyp. Nat. Struct. Mol. Biol. 16, 1036–1042 (2009)

  28. 28.

    et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. D 67, 235–242 (2011)

  29. 29.

    & Processing diffraction data with MOSFLM. Nato Sci Ser Ii Math 245, 41–51 (2007)

  30. 30.

    An introduction to data reduction: space-group determination, scaling and intensity statistics. Acta Crystallogr. D 67, 282–292 (2011)

  31. 31.

    & How good are my data and what is the resolution? Acta Crystallogr. D 69, 1204–1214 (2013)

  32. 32.

    et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007)

  33. 33.

    , & Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D 53, 240–255 (1997)

  34. 34.

    & Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004)

  35. 35.

    et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010)

  36. 36.

    , , , & Electrostatics of nanosystems: application to microtubules and the ribosome. Proc. Natl Acad. Sci. USA 98, 10037–10041 (2001)

  37. 37.

    & 3V: cavity, channel and cleft volume calculator and extractor. Nucleic Acids Res. 38, W555–W562 (2010)

  38. 38.

    et al. Mature HIV-1 capsid structure by cryo-electron microscopy and all-atom molecular dynamics. Nature 497, 643–646 (2013)

  39. 39.

    et al. Structural virology. X-ray crystal structures of native HIV-1 capsid protein reveal conformational variability. Science 349, 99–103 (2015)

  40. 40.

    , , & Replication of phenotypically mixed human immunodeficiency virus type 1 virions containing catalytically active and catalytically inactive reverse transcriptase. J. Virol. 75, 6537–6546 (2001)

Download references


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.


  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


  1. Search for David A. Jacques in:

  2. Search for William A. McEwan in:

  3. Search for Laura Hilditch in:

  4. Search for Amanda J. Price in:

  5. Search for Greg J. Towers in:

  6. Search for Leo C. James in:


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.


  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

Publication history






Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.