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Retroviral intasome assembly and inhibition of DNA strand transfer

Abstract

Integrase is an essential retroviral enzyme that binds both termini of linear viral DNA and inserts them into a host cell chromosome. The structure of full-length retroviral integrase, either separately or in complex with DNA, has been lacking. Furthermore, although clinically useful inhibitors of HIV integrase have been developed, their mechanism of action remains speculative. Here we present a crystal structure of full-length integrase from the prototype foamy virus in complex with its cognate DNA. The structure shows the organization of the retroviral intasome comprising an integrase tetramer tightly associated with a pair of viral DNA ends. All three canonical integrase structural domains are involved in extensive protein–DNA and protein–protein interactions. The binding of strand-transfer inhibitors displaces the reactive viral DNA end from the active site, disarming the viral nucleoprotein complex. Our findings define the structural basis of retroviral DNA integration, and will allow modelling of the HIV-1 intasome to aid in the development of antiretroviral drugs.

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Figure 1: Architecture of the PFV intasome.
Figure 2: Sequence-specific protein–DNA interactions.
Figure 3: PFV IN active site in committed and drug-bound states.
Figure 4: Predicted target DNA binding orientation.

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Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors have been deposited with the Protein Data Bank under accession codes 3L2Q (Apo), 3L2R (Mg), 3L2S (Mn), 3L2T (Mg/MK0518), 3L2U (Mg/GS9137), 3L2V (Mn/MK0518) and 3L2W (Mn/GS9137) structures. Raw diffraction images are available on request.

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Acknowledgements

We thank F. Dyda for critical reading of the manuscript, R. Clayton and M. Cummings for the generous gift of InSTIs and helpful discussions, T. Sorensen and the staff of the I02 and I04 beamlines of the Diamond Light Source for assistance with X-ray data collection. P.C. and co-workers are funded by the UK Medical Research Council, and A.E. by the US National Institutes of Health.

Author Contributions E.V. and P.C. carried out initial trials with truncated PFV IN constructs; S.S.G. and P.C. obtained full-length PFV IN–DNA complexes, carried out crystallization screening and optimization; S.H. soaked and prepared crystals for data collection; S.H. and P.C. collected diffraction data and solved the structures; S.H. refined the final models; S.H. and S.S.G. carried out gel-filtration and activity assays; P.C., S.H. and A.E. wrote the paper.

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Correspondence to Peter Cherepanov.

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This file contains Supplementary Table 1 and Supplementary Figures 1-7 with Legends. (PDF 5518 kb)

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This movie file shows the overall architecture of the PFV intasome. (MOV 10871 kb)

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Hare, S., Gupta, S., Valkov, E. et al. Retroviral intasome assembly and inhibition of DNA strand transfer. Nature 464, 232–236 (2010). https://doi.org/10.1038/nature08784

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