Analysis of bisdioxopiperazine dexrazoxane binding to human DNA topoisomerase II alpha: decreased binding as a mechanism of drug resistance.
Analysis of bisdioxopiperazine dexrazoxane binding to human DNA topoisomerase II alpha: decreased binding as a mechanism of drug resistance.
复制标题
双二氧代哌嗪右雷佐生与人 DNA 拓扑异构酶 II α 结合的分析：结合减少是耐药机制。
DOI：
10.1021/bi034557d
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
Sehested,Maxwell
中科院分区：
文献类型：
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作者：
Renodon-Corniere,Axelle;Jensen,LarsH;Nitiss,JohnL;Jensen,PeterB;Sehested,Maxwell
拓扑异构酶II是一种ATP操作的钳，通过捕获双链DNA片段并将其通过另一个DNA分子运输来影响拓扑变化。尽管拓扑异构酶II靶向药物在肿瘤化疗中的广泛应用以及耐药性对治疗效果的影响，但这些药物与拓扑异构酶II之间的相互作用仍不清楚。为了确定双氧哌嗪右旋氮杂环己烷与拓扑异构酶II的相互作用，我们建立了一种快速凝胶过滤方法，并表征了(~3H)-右旋氮杂环烷与人拓扑异构酶IIα的结合。在脱氧核糖核酸和三磷酸腺苷存在下，右旋氮杂环己烷与人拓扑异构酶IIα结合，Kd值为23μM，化学计量比为1个药物分子。人类拓扑异构酶IIα上的各种N末端单一氨基酸替换，以前被证明具有特异性的双氧哌嗪耐药，或者完全取消了药物结合，或者导致结合效率降低。各种突变对药物结合的影响与它们在体内和体外对耐药性的影响有很好的相关性。有趣的是，人拓扑异构酶IIα活性位点的一个改变的酪氨酸突变体不能进行DNA链传递，不能与右旋氮杂环己烷结合，这与药物在拓扑异构酶II催化循环后期提出的作用机制一致。药物结合水平与Dexrazoxane耐药性之间的直接相关性与这些导致Dexrazoxane耐药突变的药物结合减少的作用机制是一致的。
Topoisomerase II is an ATP-operated clamp that effects topological changes by capturing a double stranded DNA segment and transporting it through another DNA molecule. Despite the extensive use of topoisomerase II-targeted drugs in cancer chemotherapy and the impact of drug resistance on the efficacy of treatment, much remains unknown concerning the interactions between these agents and topoisomerase II. To identify the interaction of the bisdioxopiperazine dexrazoxane (ICRF-187) with topoisomerase II, we developed a rapid gel-filtration assay and characterized the binding of (3H)-dexrazoxane to human topoisomerase II α. Dexrazoxane binds to human topoisomerase II α in the presence of DNA and ATP with an apparentKdof 23 μM and a stoichiometry of 1 drug molecule per enzyme dimer. Various N-terminal single amino acid substitutions in human topoisomerase II α that were previously shown to confer specific bisdioxopiperazine resistance either totally abolished drug binding or resulted in less efficient binding. The effect of the various mutations on drug binding correlated well with their effect on drug resistance in vivo and in vitro. Interestingly, an altered active site tyrosine mutant of human topoisomerase II α, which is incapable of carrying out DNA strand passage, was unable to bind dexrazoxane, which agrees with the drug's proposed mechanism of action late in the topoisomerase II catalytic cycle. The direct correlation between the level of drug binding and dexrazoxane resistance is consistent with a decreased drug binding mechanism of action for these dexrazoxane resistance conferring mutations.