Probing the Molecular Mechanisms of AZT Drug Resistance Mediated by HIV-1 Reverse Transcriptase Using a Transient Kinetic Analysis
| Autor: | Joseph A. Vaccaro, Eisuke Murakami, Aravind Basavapathruni, Dagny Ulrich, Karen S. Anderson, Chung K. Chu, Adrian S. Ray, Raymond F. Schinazi |
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| Rok vydání: | 2003 |
| Předmět: |
Time Factors
Anti-HIV Agents Mutant Drug resistance Biochemistry Inhibitory Concentration 50 Structure-Activity Relationship chemistry.chemical_compound Adenosine Triphosphate Drug Resistance Viral Nucleotide chemistry.chemical_classification Binding Sites Dose-Response Relationship Drug biology DNA synthesis Active site Molecular biology HIV Reverse Transcriptase Reverse transcriptase Kinetics Terminator (genetics) Models Chemical chemistry Mutation biology.protein Reverse Transcriptase Inhibitors Thymidine Zidovudine Protein Binding |
| Zdroj: | Biochemistry. 42:8831-8841 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi034435l |
| Popis: | Several hypotheses have been proposed to explain the development of resistance to the anti-HIV drug AZT. Clinical findings show that AZT resistance mutations in HIV-1 reverse transcriptase (RT) not only reduce susceptibility to thymidine analogues but may also confer multi-dideoxynucleoside resistance. In this report, we describe transient kinetic studies establishing the biochemical effects of AZT resistance mutations in HIV-1 RT on the incorporation and removal of natural and unnatural deoxynucleotides. While the physiological role remains to be elucidated, the largest biochemical difference between wild-type and AZT resistant HIV-1 RT manifested itself during ATP-mediated deoxynucleotide removal. Enhanced removal resulted from an increase in the maximum rate of chain terminator excision, suggesting that mutated residues play a role in the optimal alignment of substrates for ATP-mediated removal. The efficiency of pyrophosphorolysis was not increased by the presence of AZT resistance mutations. However, a 2-fold decrease in the extent of inhibition caused by the next correct nucleotide during pyrophosphorolytic cleavage of a D4TMP chain-terminated primer may illustrate how this mutant can utilize pyrophosphate to enhance resistance. The inability of RT to catalyze removal of a chain terminator from an RNA-RNA primer-template may show how slight changes in selectivity against AZTMP incorporation during the initiation of DNA synthesis can contribute to high-level resistance. Taken together, these results suggest that multiple modes of resistance may be conferred by these mutations. Structure-activity studies of chain terminator removal suggest that analogues that form tight interactions with residues in the RT active site may be more prone to resistance mechanisms mediated by removal. |
| Databáze: | OpenAIRE |
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