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Journal of Virology, July 2006, p. 7169-7178, Vol. 80, No. 14
0022-538X/06/$08.00+0     doi:10.1128/JVI.00322-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Hypersusceptibility to Substrate Analogs Conferred by Mutations in Human Immunodeficiency Virus Type 1 Reverse Transcriptase

Robert A. Smith,* Donovan J. Anderson, and Bradley D. Preston

Department of Pathology, University of Washington, Seattle, Washington 98195

Received 14 February 2006/ Accepted 29 April 2006

Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) contains four structural motifs (A, B, C, and D) that are conserved in polymerases from diverse organisms. Motif B interacts with the incoming nucleotide, the template strand, and key active-site residues from other motifs, suggesting that motif B is an important determinant of substrate specificity. To examine the functional role of this region, we performed "random scanning mutagenesis" of 11 motif B residues and screened replication-competent mutants for altered substrate analog sensitivity in culture. Single amino acid replacements throughout the targeted region conferred resistance to lamivudine and/or hypersusceptibility to zidovudine (AZT). Substitutions at residue Q151 increased the sensitivity of HIV-1 to multiple nucleoside analogs, and a subset of these Q151 variants was also hypersusceptible to the pyrophosphate analog phosphonoformic acid (PFA). Other AZT-hypersusceptible mutants were resistant to PFA and are therefore phenotypically similar to PFA-resistant variants selected in vitro and in infected patients. Collectively, these data show that specific amino acid replacements in motif B confer broad-spectrum hypersusceptibility to substrate analog inhibitors. Our results suggest that motif B influences RT-deoxynucleoside triphosphate interactions at multiple steps in the catalytic cycle of polymerization.

* Corresponding author. Mailing address: University of Washington, Department of Pathology, K-084 HSB, Box 357705, 1959 NE Pacific St., Seattle, WA 98195. Phone: (206) 221-5650. Fax: (206) 543-3967. E-mail: smithra{at}u.washington.edu.

Journal of Virology, July 2006, p. 7169-7178, Vol. 80, No. 14
0022-538X/06/$08.00+0     doi:10.1128/JVI.00322-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Smith, R. A., Gottlieb, G. S., Anderson, D. J., Pyrak, C. L., Preston, B. D. (2008). Human Immunodeficiency Virus Types 1 and 2 Exhibit Comparable Sensitivities to Zidovudine and Other Nucleoside Analog Inhibitors In Vitro. Antimicrob. Agents Chemother. 52: 329-332 [Abstract] [Full Text]  


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