Portugal J, Fox K R, McLean M J, Richenberg J L, Waring M J
Department of Pharmacology, University of Cambridge, UK.
Nucleic Acids Res. 1988 May 11;16(9):3655-70. doi: 10.1093/nar/16.9.3655.
The reactivity of the 160 bp tyrT DNA fragment towards diethyl pyrocarbonate (DEPC) has been investigated in the presence of bis-intercalating quinoxaline antibiotics and the synthetic depsipeptide TANDEM. At moderate concentrations of each ligand, specific purine residues (mainly adenosines) exhibit enhanced reactivity towards the probe, and several sites of enhancement appear to be related to the sequence selectivity of drug binding. Further experiments were performed with echinomycin at pH 5.5 and 4.6 to facilitate the protonation of cytosine required for formation of Hoogsteen GC base pairs. No significant increase in reactivity was observed under these conditions. Additionally, no protection of deoxyguanosine residues from methylation by dimethyl sulphate was observed in the presence of echinomycin. We conclude that the structural anomaly giving rise to drug-dependent enhanced DEPC reaction is not simply the formation of Hoogsteen base pairs adjacent to antibiotic binding sites. Nor is it due to a general unwinding of the double helix, since we show that conditions which are supposed to unwind the helix lead to a uniform increase in purine reactivity, regardless of the surrounding nucleotide sequence.
在双插入喹喔啉抗生素和合成缩肽TANDEM存在的情况下，研究了160 bp tyrT DNA片段对焦碳酸二乙酯（DEPC）的反应性。在每种配体的适度浓度下，特定的嘌呤残基（主要是腺苷）对探针表现出增强的反应性，并且几个增强位点似乎与药物结合的序列选择性有关。用棘霉素在pH 5.5和4.6条件下进行了进一步实验，以促进形成Hoogsteen GC碱基对所需的胞嘧啶质子化。在这些条件下未观察到反应性有显著增加。此外，在棘霉素存在的情况下，未观察到脱氧鸟苷残基免受硫酸二甲酯甲基化的保护作用。我们得出结论，导致药物依赖性增强的DEPC反应的结构异常不仅仅是与抗生素结合位点相邻的Hoogsteen碱基对的形成。也不是由于双螺旋的普遍解旋，因为我们表明，假定能使螺旋解旋的条件会导致嘌呤反应性均匀增加，而与周围的核苷酸序列无关。
