The investigators propose to study the mechanism of AZT resistance and suppression of AZT resistance by suppressor mutations. They well also study RT mutations that alter catalytic activity, non-nucleoside RT inhibitors, and various nucleotide analogues. They propose to characterize quantitatively the formation of conformationally active stable complex formed between HIV-1 RT, primer-template, and deoxyribonucleoside triphosphate (dNTP) when phosphodiester bond synthesis is prevented. They well monitor the formational changes that occur in the formation of stable complex by measuring nuclease protection, dissociation rate in absence of dNTP, circular dichroism spectral changes, and changes in intrinsic protein fluorescence. In addition, they will study the effects of these mutations on RNase H endonuclease and exonuclease activity. Finally, they well characterize a novel AZTTP-dependent DNA modifying activity of HIV-1 RT that they recently identified. The unifying hypothesis is that drug-resistance mutations render the enzyme less effective at DNA chain termination.

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National Institute of Allergy and Infectious Diseases (NIAID)
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AIDS and Related Research Study Section 4 (ARRD)
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University of Miami School of Medicine
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Lu, Gaofei; Matsuura, Suzanne E; Barrientos, Antoni et al. (2013) HIV-1 infection is blocked at an early stage in cells devoid of mitochondrial DNA. PLoS One 8:e78035
Acosta-Hoyos, Antonio J; Matsuura, Suzanne E; Meyer, Peter R et al. (2012) A role of template cleavage in reduced excision of chain-terminating nucleotides by human immunodeficiency virus type 1 reverse transcriptase containing the M184V mutation. J Virol 86:5122-33
Scott, Walter A (2011) Structures of reverse transcriptase pre- and post-excision complexes shed new light on HIV-1 AZT resistance. Viruses 3:20-5
Acosta-Hoyos, Antonio J; Scott, Walter A (2010) The Role of Nucleotide Excision by Reverse Transcriptase in HIV Drug Resistance. Viruses 2:372-394
Rutvisuttinunt, Wiriya; Meyer, Peter R; Scott, Walter A (2008) Interactions between HIV-1 reverse transcriptase and the downstream template strand in stable complexes with primer-template. PLoS One 3:e3561
Meyer, Peter R; Rutvisuttinunt, Wiriya; Matsuura, Suzanne E et al. (2007) Stable complexes formed by HIV-1 reverse transcriptase at distinct positions on the primer-template controlled by binding deoxynucleoside triphosphates or foscarnet. J Mol Biol 369:41-54
Meyer, Peter R; Smith, Anthony J; Matsuura, Suzanne E et al. (2006) Chain-terminating dinucleoside tetraphosphates are substrates for DNA polymerization by human immunodeficiency virus type 1 reverse transcriptase with increased activity against thymidine analogue-resistant mutants. Antimicrob Agents Chemother 50:3607-14
Smith, Anthony James; Scott, Walter Alvin (2006) The influence of natural substrates and inhibitors on the nucleotide-dependent excision activity of HIV-1 reverse transcriptase in the infected cell. Curr Pharm Des 12:1827-41
Smith, Anthony J; Meyer, Peter R; Asthana, Deshratn et al. (2005) Intracellular substrates for the primer-unblocking reaction by human immunodeficiency virus type 1 reverse transcriptase: detection and quantitation in extracts from quiescent- and activated-lymphocyte subpopulations. Antimicrob Agents Chemother 49:1761-9
Meyer, Peter R; Smith, Anthony J; Matsuura, Suzanne E et al. (2004) Effects of primer-template sequence on ATP-dependent removal of chain-terminating nucleotide analogues by HIV-1 reverse transcriptase. J Biol Chem 279:45389-98

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