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Structure of the immature retroviral capsid at 8 Å resolution by cryo-electron microscopy

Nature volume 487, pages 385–389 (2012)Cite this article

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Abstract

The assembly of retroviruses such as HIV-1 is driven by oligomerization of their major structural protein, Gag. Gag is a multidomain polyprotein including three conserved folded domains: MA (matrix), CA (capsid) and NC (nucleocapsid)1. Assembly of an infectious virion proceeds in two stages2. In the first stage, Gag oligomerization into a hexameric protein lattice leads to the formation of an incomplete, roughly spherical protein shell that buds through the plasma membrane of the infected cell to release an enveloped immature virus particle. In the second stage, cleavage of Gag by the viral protease leads to rearrangement of the particle interior, converting the non-infectious immature virus particle into a mature infectious virion. The immature Gag shell acts as the pivotal intermediate in assembly and is a potential target for anti-retroviral drugs both in inhibiting virus assembly and in disrupting virus maturation3. However, detailed structural information on the immature Gag shell has not previously been available. For this reason it is unclear what protein conformations and interfaces mediate the interactions between domains and therefore the assembly of retrovirus particles, and what structural transitions are associated with retrovirus maturation. Here we solve the structure of the immature retroviral Gag shell from Mason–Pfizer monkey virus by combining cryo-electron microscopy and tomography. The 8-Å resolution structure permits the derivation of a pseudo-atomic model of CA in the immature retrovirus, which defines the protein interfaces mediating retrovirus assembly. We show that transition of an immature retrovirus into its mature infectious form involves marked rotations and translations of CA domains, that the roles of the amino-terminal and carboxy-terminal domains of CA in assembling the immature and mature hexameric lattices are exchanged, and that the CA interactions that stabilize the immature and mature viruses are almost completely distinct.

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Figure 1: Cryo-EM reconstruction of M-PMV CANC tubes.
Figure 2: Fitting of known atomic structures into the cryo-EM map.
Figure 3: Comparison of the arrangement of CA domains in the mature and immature retrovirus lattices.
Figure 4: Modularity of protein interfaces.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Cryo-EM structural data have been deposited at the EMDB under accession numbers EMD-2089 and EMD-2090, and at the Protein Data Bank under accession numbers 4ard and 4arg.

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Acknowledgements

This study was technically supported by the use of the European Molecular Biology Laboratory Information Technology Service unit. This work was partly funded by a grant from the Deutsche Forschungsgemeinschaft within SPP1175 to J.A.G.B. and by grants P302/12/1895 and 204/09/1388 from the Czech Science foundation to T.R. and M.R.

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Author notes

  1. Norman E. Davey and Pavel Ulbrich: These authors contributed equally to this work.

Authors and Affiliations

  1. Structural and Computational Biology Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany,

    Tanmay A. M. Bharat, Norman E. Davey, James D. Riches, Alex de Marco, Carsten Sachse & John A. G. Briggs

  2. Department of Biochemistry and Microbiology, Institute of Chemical Technology, Prague, Technicka 3, 166 28 Prague, Czech Republic,

    Pavel Ulbrich & Tomas Ruml

  3. Institute of Organic Chemistry and Biochemistry, Academy of Sciences of Czech Republic, v.v.i., Flemingovo nám. 2, 166 10 Prague, Czech Republic,

    Michaela Rumlova

Authors
  1. Tanmay A. M. Bharat
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Contributions

T.A.M.B., P.U., M.R., T.R. and J.A.G.B. designed the research. T.A.M.B. and P.U. prepared samples for electron microscopy. T.A.M.B. and J.D.R. collected cryo-EM data. T.A.M.B., J.D.R., A.D.M. and J.A.G.B. analysed cryo-ET data. C.S. supported helical image-processing techniques. T.A.M.B. and J.A.G.B. developed and applied the variable-symmetry helical reconstruction methodology. T.A.M.B., N.D., P.U., M.R., C.S., T.R. and J.A.G.B. analysed fitted pseudo-atomic models. T.A.M.B. and J.A.G.B. wrote the paper with support from all the authors.

Corresponding author

Correspondence to John A. G. Briggs.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-8, Supplementary Methods, Supplementary Tables 1-2, legends for Supplementary Movies 1-2 and Supplementary References. (PDF 3688 kb)

Supplementary Movie 1

In this movie we see the structure of the M-PMV CANC tubes at sub-nanometre resolution - see Supplementary Information file for full legend. (MOV 16313 kb)

Supplementary Movie 2

In this movie we see the comparison of the immature and mature retroviral lattices - see Supplementary Information file for full legend. (MOV 5283 kb)

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Bharat, T., Davey, N., Ulbrich, P. et al. Structure of the immature retroviral capsid at 8 Å resolution by cryo-electron microscopy. Nature 487, 385–389 (2012). https://doi.org/10.1038/nature11169

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