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Proc Natl Acad Sci U S A. 2019 Apr 9;116(15):7308-7313. doi: 10.1073/pnas.1814170116. Epub 2019 Mar 22.

Structure of HIV-1 RT/dsRNA initiation complex prior to nucleotide incorporation.

Das K1,2,3,4, Martinez SE5,2,3,4, DeStefano JJ6,7, Arnold E1,2.

Author information

1
Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, NJ 08854; kalyan.das@kuleuven.be arnold@cabm.rutgers.edu.
2
Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854.
3
Rega Institute, The Katholieke Universiteit Leuven, 3000 Leuven, Belgium.
4
Department of Microbiology, Immunology and Transplantation, The Katholieke Universiteit Leuven, 3000 Leuven, Belgium.
5
Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, NJ 08854.
6
Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742.
7
Maryland Pathogen Research Institute, University of Maryland, College Park, MD 20742.

Abstract

The initiation phase of HIV reverse transcription has features that are distinct from its elongation phase. The first structure of a reverse transcription initiation complex (RTIC) that trapped the complex after incorporation of one ddCMP nucleotide was published recently [Larsen KP, et al. (2018) Nature 557:118-122]. Here we report a crystal structure of a catalytically active HIV-1 RT/dsRNA complex that mimics the state of the RTIC before the first nucleotide incorporation. The structure reveals that the dsRNA-bound conformation of RT is closer to that of RT bound to a nonnucleoside RT inhibitor (NNRTI) and dsDNA; a hyperextended thumb conformation helps to accommodate the relatively wide dsRNA duplex. The RNA primer 3' end is positioned 5 Å away from the polymerase site; however, unlike in an NNRTI-bound state in which structural elements of RT restrict the movement of the primer, the primer terminus of dsRNA is not blocked from reaching the active site of RT. The observed structural changes and energetic cost of bringing the primer 3' end to the priming site are hypothesized to explain the slower nucleotide incorporation rate of the RTIC. An unusual crystal lattice interaction of dsRNA with its symmetry mate is reminiscent of the RNA architecture within the extended vRNA-tRNALys3 in the RTIC. This RT/dsRNA complex captures the key structural characteristics and components of the RTIC, including the RT conformational changes and interactions with the dsRNA primer-binding site region, and these features have implications for better understanding of RT initiation.

KEYWORDS:

DNA polymerase; HIV reverse transcriptase; RTIC; double-stranded RNA structure; reverse transcription initiation

PMID:
30902895
PMCID:
PMC6462067
DOI:
10.1073/pnas.1814170116
[Indexed for MEDLINE]
Free PMC Article

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