Article

Nature 453, 184-189 (8 May 2008) | doi:10.1038/nature06941; Received 12 November 2007; Accepted 20 March 2008

Dynamic binding orientations direct activity of HIV reverse transcriptase

Elio A. Abbondanzieri1,5, Gregory Bokinsky1,5, Jason W. Rausch4, Jennifer X. Zhang1, Stuart F. J. Le Grice4 & Xiaowei Zhuang1,2,3

  1. Department of Chemistry and Chemical Biology,
  2. Department of Physics, and,
  3. Howard Hughes Medical Institute, Harvard University, Cambridge, Massachusetts 02138, USA
  4. HIV Drug Resistance Program, National Cancer Institute, Frederick, Maryland 21702, USA
  5. These authors contributed equally to this work.

Correspondence to: Xiaowei Zhuang1,2,3 Correspondence and requests for materials should be addressed to X.Z. (Email: zhuang@chemistry.harvard.edu).

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The reverse transcriptase of human immunodeficiency virus (HIV) catalyses a series of reactions to convert the single-stranded RNA genome of HIV into double-stranded DNA for host-cell integration. This task requires the reverse transcriptase to discriminate a variety of nucleic-acid substrates such that active sites of the enzyme are correctly positioned to support one of three catalytic functions: RNA-directed DNA synthesis, DNA-directed DNA synthesis and DNA-directed RNA hydrolysis. However, the mechanism by which substrates regulate reverse transcriptase activities remains unclear. Here we report distinct orientational dynamics of reverse transcriptase observed on different substrates with a single-molecule assay. The enzyme adopted opposite binding orientations on duplexes containing DNA or RNA primers, directing its DNA synthesis or RNA hydrolysis activity, respectively. On duplexes containing the unique polypurine RNA primers for plus-strand DNA synthesis, the enzyme can rapidly switch between the two orientations. The switching kinetics were regulated by cognate nucleotides and non-nucleoside reverse transcriptase inhibitors, a major class of anti-HIV drugs. These results indicate that the activities of reverse transcriptase are determined by its binding orientation on substrates.

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