The different cleavage DNA sequence specificity explains the camptothecin resistance of the human topoisomerase I Glu418Lys mutant.-猫眼课题宝

DOI：

10.1093/nar/gkl670

发表时间：

2006

期刊：

Nucleic acids research

影响因子：

14.9

通讯作者：

中科院分区：

生物学2区

文献类型：

作者：

Fiorani P;Chillemi G;Losasso C;Castelli S;Desideri A

来源链接：

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1636438/?report=classic

表达Glu 418 Lys人拓扑异构酶I突变体的酵母细胞显示出作为时间的函数缓慢降低的喜树碱抗性。纯化的突变体的催化循环的单个步骤的分子表征表明，它具有与野生型蛋白相同的松弛活性，但与野生型酶相比，对切割位点具有不同的DNA序列特异性，如在几种底物上测定的。特别地，突变体对CPT敏感的可切割位点具有低特异性。事实上，与野生型酶不同，突变体对易切割链-1位有胸腺嘧啶碱基的切割位点的偏好降低。这种偏好，加上对有效结合喜树碱的-1位胸腺嘧啶碱基的严格要求，解释了表达突变体的酵母细胞对喜树碱的暂时抗性，并指出了DNA序列在喜树碱药物结合中的重要性。

Yeast cells expressing the Glu418Lys human topoisomerase I mutant display a camptothecin resistance that slowly decreases as a function of time. Molecular characterization of the single steps of the catalytic cycle of the purified mutant indicates that it has a relaxation activity identical to the wild-type protein but a different DNA sequence specificity for the cleavage sites when compared to the wild-type enzyme, as assayed on several substrates. In particular the mutant has a low specificity for CPT sensitive cleavable sites. In fact, the mutant has, at variance of the wild-type enzyme, a reduced preference for cleavage sites having a thymine base in position −1 of the scissile strand. This preference, together with the strict requirement for a thymine base in position −1 for an efficient camptothecin binding, explains the temporary camptothecin resistance of the yeast cell expressing the mutant and points out the importance of the DNA sequence in the binding of the camptothecin drug.