3“-azido-3”脱氧胸苷磷酸化和选择性交互的5 '三磷酸与人类免疫缺陷病毒逆转录酶。
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福曼PA, Fyfe是的,圣克莱尔MH Weinhold K,赖德奥特莱托,弗里曼GA,理SN, Bolognesi DP,布罗德年代,迁址H, et al。
3“-azido-3”脱氧胸苷磷酸化和选择性交互的5 '三磷酸与人类免疫缺陷病毒逆转录酶。
《美国国家科学院刊S a . 1986年11月,83 (21):8333 - 7。
- PubMed ID
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2430286 (在PubMed]
- 文摘
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胸苷模拟3 ' -azido-3脱氧胸苷(BW A509U叠氮胸苷)能有效地抑制人类免疫缺陷病毒(HIV)复制在50 - 500纳米范围内(迁址,H。Weinhold, k . J。福曼,p。圣克莱尔,m . H。、Nusinoff-Lehrman年代。盖洛,r . C。Bolognesi D。巴里,d . w . &布罗德,美国(1985年)Proc。国家的。学会科学。美国82年,7096 - 7100]。相比之下,抑制感染人类成纤维细胞和淋巴细胞的生长已经观察到只有在浓度超过1毫米。这种选择性是调查的性质。叠氮胸苷合成代谢的5 ' mono - di -,三磷酸衍生物是相似的感染和艾滋病毒感染细胞。叠氮胸苷一磷酸的水平很高,而di -三磷酸的水平较低(小于或等于5 microM和小于或等于2 microM,分别)。 Cytosolic thymidine kinase (EC 2.7.1.21) was responsible for phosphorylation of azidothymidine to its monophosphate. Purified thymidine kinase catalyzed the phosphorylations of thymidine and azidothymidine with apparent Km values of 2.9 microM and 3.0 microM. The maximal rate of phosphorylation with azidothymidine was equal to 60% of the rate with thymidine. Phosphorylation of azidothymidine monophosphate to the diphosphate also appeared to be catalyzed by a host-cell enzyme, thymidylate kinase (EC 2.7.4.9). The apparent Km value for azidothymidine monophosphate was 2-fold greater than the value for dTMP (8.6 microM vs. 4.1 microM), but the maximal phosphorylation rate was only 0.3% of the dTMP rate. These kinetic constants were consistent with the anabolism results and indicated that azidothymidine monophosphate is an alternative-substrate inhibitor of thymidylate kinase. This conclusion was reflected in the observation that cells incubated with azidothymidine had reduced intracellular levels of dTTP. IC50 (concentration of inhibitor that inhibits enzyme activity 50%) values were determined for azidothymidine triphosphate with HIV reverse transcriptase and with immortalized human lymphocyte (H9 cell) DNA polymerase alpha. Azidothymidine triphosphate competed about 100-fold better for the HIV reverse transcriptase than for the cellular DNA polymerase alpha. The results reported here suggest that azidothymidine is nonselectively phosphorylated but that the triphosphate derivative efficiently and selectively binds to the HIV reverse transcriptase. Incorporation of azidothymidylate into a growing DNA strand should terminate DNA elongation and thus inhibit DNA synthesis.