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The mechanism of retroviral integration from X-ray structures of its key intermediates

Abstract

To establish productive infection, a retrovirus must insert a DNA replica of its genome into host cell chromosomal DNA1,2. This process is operated by the intasome, a nucleoprotein complex composed of an integrase tetramer (IN) assembled on the viral DNA ends3,4. The intasome engages chromosomal DNA within a target capture complex to carry out strand transfer, irreversibly joining the viral and cellular DNA molecules. Although several intasome/transpososome structures from the DDE(D) recombinase superfamily have been reported4,5,6, the mechanics of target DNA capture and strand transfer by these enzymes remained unclear. Here we report crystal structures of the intasome from prototype foamy virus in complex with target DNA, elucidating the pre-integration target DNA capture and post-catalytic strand transfer intermediates of the retroviral integration process. The cleft between IN dimers within the intasome accommodates chromosomal DNA in a severely bent conformation, allowing widely spaced IN active sites to access the scissile phosphodiester bonds. Our results resolve the structural basis for retroviral DNA integration and provide a framework for the design of INs with altered target sequences.

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Figure 1: Crystal structure of the PFV STC.
Figure 2: Details of DNA conformations, recognition, and active site mechanics during strand transfer.
Figure 3: Sequence analysis of strand transfer reaction products.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors for STC, TCCddA and TCCApo have been deposited with the Protein Data Bank under accession codes 3OS0, 3OS1 and 3OS2, respectively. Raw diffraction images are available upon request.

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Acknowledgements

We thank A. Engelman and F. Dyda for critical reading of the manuscript, J. Sanchez-Weatherby for assistance with X-ray data collection at the I02 beamline of the Diamond Light Source and J. Moore for help with crystallization screening and the X-ray generator. This work was funded by the UK Medical Research Council.

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Authors and Affiliations

Authors

Contributions

G.N.M., S.H. and P.C. performed the experiments and wrote the paper.

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Correspondence to Peter Cherepanov.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

The file contains Supplementary Table 1 and Supplementary Figures 1-8 with legends. (PDF 1224 kb)

Supplementary Movie 1

This movie shows the overall architecture of the PFV STC. Protein and DNA chains are coloured as in Fig. 1b and c. (MOV 8200 kb)

Supplementary Movie 2

This movie illustrates tDNA bending that transpires when the intasome engages hypothetical initial B-form tDNA to form the TCC and the subsequent DNA strand transfer. For clarity, only DNA strands are shown. (MOV 3119 kb)

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Maertens, G., Hare, S. & Cherepanov, P. The mechanism of retroviral integration from X-ray structures of its key intermediates. Nature 468, 326–329 (2010). https://doi.org/10.1038/nature09517

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