Kjeldsen E, Mollerup S, Thomsen B, Bonven B J, Bolund L, Westergaard O
Department of Molecular Biology and Plant Physiology, University of Aarhus, Denmark.
J Mol Biol. 1988 Jul 20;202(2):333-42. doi: 10.1016/0022-2836(88)90462-7.
We have studied the effect of the antitumor drug, camptothecin, on the interaction of human topoisomerase I with DNA at the sequence level. At a low molar ratio of enzyme to DNA, cleavage is prominent and unique, located at a previously described hexadecameric recognition sequence, while a number of strong additional cleavage sites appear in the presence of the drug. Camptothecin stimulates cleavage at the recognition sequence less than twofold, whereas cleavage at the additional sites is stimulated up to 200-fold. Camptothecin greatly enhances the stability of the cleavable complexes formed at the additional sites, whereas the complex formed at the hexadecameric sequence is only marginally affected. Cleavage was eliminated at certain sites in the presence of camptothecin. Taken together these observations demonstrate that at least three types of potential eukaryotic topoisomerase I cleavage sites can be distinguished by the use of camptothecin. Comparison of the sequences at the additional cleavage sites in the presence of camptothecin reveals that the most frequently cleaved dinucleotide is TG with no consensus for the flanking nucleotides.
我们已经在序列水平上研究了抗肿瘤药物喜树碱对人拓扑异构酶I与DNA相互作用的影响。在酶与DNA的低摩尔比下，切割作用显著且独特，位于先前描述的十六聚体识别序列处，而在药物存在的情况下会出现许多额外的强切割位点。喜树碱刺激识别序列处的切割作用不到两倍，而在额外位点处的切割作用则被刺激高达200倍。喜树碱极大地增强了在额外位点形成的可切割复合物的稳定性，而在十六聚体序列处形成的复合物仅受到轻微影响。在喜树碱存在的情况下，某些位点的切割作用被消除。综合这些观察结果表明，通过使用喜树碱可以区分至少三种潜在的真核生物拓扑异构酶I切割位点。对喜树碱存在时额外切割位点处的序列进行比较发现，最常被切割的二核苷酸是TG，其侧翼核苷酸没有一致序列。
