Shelton C J, Harding M M, Prakash A S
School of Chemistry, University of Sydney, N.S.W., Australia.
Biochemistry. 1996 Jun 18;35(24):7974-82. doi: 10.1021/bi952495o.
DNase I and three DNA chemical footprinting agents were used to compare the DNA binding properties of the anthracycline antitumor antibiotics daunomycin, aclacinomycin A, and ditrisarubicin B. These anthracyclines contain a tetracyclic chromophore which intercalates into DNA and a monosaccharide, trisaccharide, and two trisaccharide side chains, respectively. These side chains consist of between one and three 2,6-dideoxy, 1,4-diaxially linked sugars. Three chemical probes, fotemustine, dimethyl sulfate, 4-(2'-bromoethyl)phenol, and the enzymic probe DNase I were used in the footprinting experiments. The chemical probes provided a clear picture of the binding pattern at 37 degrees C and more detailed information than that obtained using the standard DNase I footprinting assay. All three anthracyclines showed preferred binding to 5'-GT-3' sequences in both the chemical and enzymatic footprinting. DNase I footprinting showed that the number of base pairs of DNA protected from cleavage increased with the number of saccharide groups present at particular sites and is consistent with DNA binding of the saccharide side chains. Alkylation of runs of guanine by fotemustine was inhibited by all three anthracyclines, while alkylation by dimethyl sulfate was enhanced for most guanines. The probe 4-(2'-bromoethyl)phenol showed that all three anthracyclines completely protected all of the adenines in the minor groove from alkylation, and enhanced major groove guanine alkylation was observed with aclacinomycin A, daunomycin, and, to a much lesser extent, ditrisarubicin B. These results are consistent with intercalation of the aglycone ring and binding of the rigid, hydrophobic saccharide side chains in the minor groove. Footprinting of four methyl glycosides related to the anthracyclines showed no evidence of DNA binding with any of the agents studied.
使用脱氧核糖核酸酶I（DNase I）和三种DNA化学足迹试剂，比较了蒽环类抗肿瘤抗生素柔红霉素、阿克拉霉素A和双三糖阿霉素B与DNA的结合特性。这些蒽环类药物含有一个嵌入DNA的四环发色团以及分别为单糖、三糖和两个三糖侧链。这些侧链由一至三个2,6 - 二脱氧、1,4 - 双轴连接的糖组成。在足迹实验中使用了三种化学探针福莫司汀、硫酸二甲酯、4 - （2'-溴乙基）苯酚以及酶促探针DNase I。化学探针清晰显示了37摄氏度时的结合模式，且比使用标准DNase I足迹分析获得的信息更详细。在化学和酶促足迹实验中，所有三种蒽环类药物均显示出对5'-GT-3'序列的优先结合。DNase I足迹分析表明，免受切割的DNA碱基对数随着特定位点存在的糖基团数量增加而增加，这与糖侧链与DNA的结合情况一致。福莫司汀对鸟嘌呤序列的烷基化作用受到所有三种蒽环类药物的抑制，而硫酸二甲酯对大多数鸟嘌呤的烷基化作用增强。探针4 - （2'-溴乙基）苯酚表明，所有三种蒽环类药物完全保护了小沟中的所有腺嘌呤不被烷基化，并且观察到阿克拉霉素A、柔红霉素以及程度小得多的双三糖阿霉素B增强了大沟鸟嘌呤的烷基化。这些结果与糖苷配基环的嵌入以及刚性疏水糖侧链在小沟中的结合情况一致。对与蒽环类药物相关的四种甲基糖苷进行的足迹分析表明，在所研究的任何试剂中均未发现与DNA结合的证据。
