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Structural basis of HIV-1 resistance to AZT by excision

Nature Structural & Molecular Biology volume 17, pages 12021209 (2010) | Download Citation

Abstract

Human immunodeficiency virus (HIV-1) develops resistance to 3′-azido-2′,3′-deoxythymidine (AZT, zidovudine) by acquiring mutations in reverse transcriptase that enhance the ATP-mediated excision of AZT monophosphate from the 3′ end of the primer. The excision reaction occurs at the dNTP-binding site, uses ATP as a pyrophosphate donor, unblocks the primer terminus and allows reverse transcriptase to continue viral DNA synthesis. The excision product is AZT adenosine dinucleoside tetraphosphate (AZTppppA). We determined five crystal structures: wild-type reverse transcriptase–double-stranded DNA (RT–dsDNA)–AZTppppA; AZT-resistant (AZTr; M41L D67N K70R T215Y K219Q) RT–dsDNA–AZTppppA; AZTr RT–dsDNA terminated with AZT at dNTP- and primer-binding sites; and AZTr apo reverse transcriptase. The AMP part of AZTppppA bound differently to wild-type and AZTr reverse transcriptases, whereas the AZT triphosphate part bound the two enzymes similarly. Thus, the resistance mutations create a high-affinity ATP-binding site. The structure of the site provides an opportunity to design inhibitors of AZT-monophosphate excision.

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Acknowledgements

We acknowledge personnel at the Cornell High Energy Synchrotron Source and the Advanced Photon Source for support of data collection, the members of our laboratories, including R. Bandwar and S. Martinez, for valuable conversations and assistance, and P. Clark for assistance with protein preparation. We are grateful to the US National Institutes of Health (NIH; grants R37 MERIT Award AI 27690 to E.A. and P01 GM 066671 to E.A. and R.A.J.) for support of reverse transcriptase structural studies. S.H.H. was supported by the Intramural Research Program of NIH, US National Cancer Institute (NCI), Center for Cancer Research and US National Institute of General Medical Sciences. This research was supported, in part, by the Intramural Research Program of the NIH, NCI, Center for Cancer Research.

Author information

Author notes

    • Stefan G Sarafianos

    Present address: Department of Molecular Microbiology & Immunology, University of Missouri, Columbia, Missouri, USA.

    • Xiongying Tu
    •  & Kalyan Das

    These authors contributed equally to this work.

Affiliations

  1. Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, USA.

    • Xiongying Tu
    • , Kalyan Das
    • , Joseph D Bauman
    • , Arthur D Clark Jr
    • , Yulia V Frenkel
    • , Stefan G Sarafianos
    •  & Eddy Arnold
  2. Department of Chemistry and Biology, Rutgers University, Piscataway, New Jersey, USA.

    • Xiongying Tu
    • , Kalyan Das
    • , Qianwei Han
    • , Joseph D Bauman
    • , Arthur D Clark Jr
    • , Xiaorong Hou
    • , Yulia V Frenkel
    • , Barbara L Gaffney
    • , Roger A Jones
    • , Stefan G Sarafianos
    •  & Eddy Arnold
  3. HIV Drug Resistance Program, National Cancer Institute–Frederick, Frederick, Maryland, USA.

    • Paul L Boyer
    •  & Stephen H Hughes

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Contributions

X.T. carried out the research, did analysis and helped with writing the manuscript. K.D. advised on crystallographic studies and did analysis, writing and editing of the manuscript. Q.H., J.D.B., X.H., B.L.G. and R.A.J. contributed special reagents. A.D.C. and Y.V.F. carried out parts of the research. P.L.B. contributed special reagents and manuscript editing. S.H.H. carried out analysis and manuscript writing and editing. S.G.S. designed the research and advised on experiments and manuscript editing. E.A. designed the research, supervised the project and did analysis, writing and editing of the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Eddy Arnold.

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https://doi.org/10.1038/nsmb.1908

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