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Mechanism of polypurine tract primer generation by HIV-1 reverse transcriptase

  1. 1.
    SYSNO ASEP0486050
    Document TypeJ - Journal Article
    R&D Document TypeJournal Article
    Subsidiary JČlánek ve WOS
    TitleMechanism of polypurine tract primer generation by HIV-1 reverse transcriptase
    Author(s) Figiel, M. (PL)
    Krepl, Miroslav (BFU-R) ORCID
    Park, S. (US)
    Poznanski, J. (PL)
    Skowronek, K. (PL)
    Golab, A. (PL)
    Ha, T. (US)
    Šponer, Jiří (BFU-R) RID, ORCID
    Nowotny, M. (PL)
    Number of authors9
    Source TitleJournal of Biological Chemistry. - : Elsevier - ISSN 0021-9258
    Roč. 293, č. 1 (2018), s. 191-202
    Number of pages12 s.
    Publication formPrint - P
    Languageeng - English
    CountryUS - United States
    Keywordshuman-immunodeficiency-virus ; strand dna-synthesis ; retroviral rnases-h ; rna/dna hybrid
    Subject RIVCE - Biochemistry
    OECD categoryBiochemistry and molecular biology
    R&D ProjectsGBP305/12/G034 GA ČR - Czech Science Foundation (CSF)
    Institutional supportBFU-R - RVO:68081707
    UT WOS000419453200017
    DOI10.1074/jbc.M117.798256
    AnnotationHIV-1 reverse transcriptase (RT) possesses both DNA polymerase activity and RNase H activity that act in concert to convert single-stranded RNA of the viral genome to double-stranded DNA that is then integrated into the DNA of the infected cell. Reverse transcriptase-catalyzed reverse transcription critically relies on the proper generation of a polypurine tract (PPT) primer. However, the mechanism of PPT primer generation and the features of the PPT sequence that are critical for its recognition by HIV-1 RT remain unclear. Here, we used a chemical cross-linking method together with molecular dynamics simulations and single-molecule assays to study the mechanism of PPT primer generation. We found that the PPT was specifically and properly recognized within covalently tethered HIV-1 RT-nucleic acid complexes. These findings indicated that recognition of the PPT occurs within a stable catalytic complex after its formation. We found that this unique recognition is based on two complementary elements that rely on the PPT sequence: RNase H sequence preference and incompatibility of the poly(rA/dT) tract of the PPT with the nucleic acid conformation that is required for RNase H cleavage. The latter results from rigidity of the poly(rA/dT) tract and leads to base-pair slip-page of this sequence upon deformation into a catalytically relevant geometry. In summary, our results reveal an unexpected mechanism of PPT primer generation based on specific dynamic properties of the poly(rA/dT) segment and help advance our understanding of the mechanisms in viral RNA reverse transcription.
    WorkplaceInstitute of Biophysics
    ContactJana Poláková, polakova@ibp.cz, Tel.: 541 517 244
    Year of Publishing2018
Number of the records: 1  

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