Mao Q, Fulmer P, Li W, DeRose E F, Petering D H
Department of Chemistry, University of Wisconsin, Milwaukee 53201, USA.
J Biol Chem. 1996 Mar 15;271(11):6185-91. doi: 10.1074/jbc.271.11.6185.
Conformational properties of HO2(-)-Co(III)-bleomycin A2 (Form I) and Co(III)-bleomycin (Form II) bound to DNA oligomers offering either principal cleavage site for the drug, d(GGAAGCTTCC)2 or d(AAACGTIT)2, have been studied by NMR methods. Form I binds in slow exchange to these oligomers. It retains most of its solution nuclear Overhauser effects (NOEs) upon binding to either oligomer. Pyrimidinyl methyl protons from the metal domain of the drug make an NOE connection with a G5 2-amino proton on DNA. The bithiazole intercalates between base pairs involving either C6 and T7 or T6 and T7 of the two DNA molecules, according to NOE connections between the bithiazole protons and protons from these bases and changes in the positions of their chemical shifts. Form II also retains most of its solution NOEs upon association with the first oligomer. However, in contrast to Form I it binds to DNA in fast exchange on the NMR time scale over the temperature range of 5-35 degrees C and does not break the degeneracy of the DNA proton chemical shifts. No intermolecular NOEs between Form II and the 10-mer have been detected. Likewise, the major perturbation in chemical shift of the histidine H2 and guanine G5 protons seen in Form I-DNA adducts is absent in Form II-DNA. The association constant of Form II with d(GGAAGCTTCC)2 in 20 mM HEPES buffer at pH 7.4 and 25 degrees C is 1.7 x 10(5) M(-1), and 1.0 mol of Form II bind per mol of 10-mer.
已通过核磁共振方法研究了与为药物提供主要切割位点的DNA寡聚物d(GGAAGCTTCC)2或d(AAACGTIT)2结合的HO2(-)-Co(III)-博来霉素A2（形式I）和Co(III)-博来霉素（形式II）的构象性质。形式I与这些寡聚物以慢交换方式结合。与任一寡聚物结合后，它保留了大部分溶液中的核Overhauser效应（NOE）。药物金属结构域的嘧啶基甲基质子与DNA上的G5 2-氨基质子形成NOE连接。根据双噻唑质子与这些碱基的质子之间的NOE连接以及它们化学位移位置的变化，双噻唑插入两个DNA分子中涉及C6和T7或T6和T7的碱基对之间。形式II与第一种寡聚物缔合后也保留了大部分溶液中的NOE。然而，与形式I不同的是，在5至35摄氏度的温度范围内，它在核磁共振时间尺度上与DNA以快交换方式结合，并且不会打破DNA质子化学位移的简并性。未检测到形式II与10聚体之间的分子间NOE。同样，在形式II-DNA中不存在形式I-DNA加合物中观察到的组氨酸H2和鸟嘌呤G5质子化学位移的主要扰动。在pH 7.4和25摄氏度的20 mM HEPES缓冲液中，形式II与d(GGAAGCTTCC)2的缔合常数为1.7 x 10(5) M(-1)，每摩尔10聚体结合1.0摩尔形式II。
