Gonzaga P E, Harris L, Margison G P, Brent T P
Department of Biochemical and Clinical Pharmacology, St Jude Children's Research Hospital, Memphis, TN 38101.
Nucleic Acids Res. 1990 Jul 11;18(13):3961-6. doi: 10.1093/nar/18.13.3961.
Chloroethylnitrosoureas (CENUs) are thought to induce cytotoxic DNA interstrand cross-links via an initial reaction at O6-position of guanine, yielding a rearranged intermediate, O6,N1-ethanoguanine. Repair of these adducts by mammalian and bacterial DNA alkyltransferases blocks the formation of cross-links. Human alkyltransferase can form a covalent complex with DNA containing BCNU-induced cross-link precursors, but the nature of the DNA-protein linkage remains unknown. Using E. coli alkyltransferases expressed by the ada and ogt genes, we now demonstrate that both enzymes can form such complexes with CENU-treated DNA. We attribute this reaction to the O6-alkylguanine repair function, because an N-terminal fragment of the ada protein, which has only alkylphosphotriester repair activity, failed to form a similar complex. This result is consistent with the idea that complex formation requires an alkyltransferase reaction with a guanine adduct, such as O6,N1-ethanoguanine. It tends to exclude the possibility that such reactions simply involve alkylation of the enzyme by reactive DNA adducts such as chloroethylphosphate or chloroethylguanine.
氯乙基亚硝脲（CENUs）被认为是通过在鸟嘌呤的O6位发生初始反应来诱导细胞毒性DNA链间交联，产生重排中间体O6，N1 - 乙撑鸟嘌呤。哺乳动物和细菌的DNA烷基转移酶对这些加合物的修复会阻止交联的形成。人类烷基转移酶可以与含有卡氮芥诱导的交联前体的DNA形成共价复合物，但DNA - 蛋白质连接的性质仍然未知。利用由ada和ogt基因表达的大肠杆菌烷基转移酶，我们现在证明这两种酶都可以与经CENU处理的DNA形成这样的复合物。我们将此反应归因于O6 - 烷基鸟嘌呤修复功能，因为ada蛋白的N端片段仅具有烷基磷酸三酯修复活性，未能形成类似的复合物。这一结果与复合物形成需要烷基转移酶与鸟嘌呤加合物（如O6，N1 - 乙撑鸟嘌呤）发生反应的观点一致。它倾向于排除此类反应仅仅涉及酶被诸如氯乙基磷酸酯或氯乙基鸟嘌呤等反应性DNA加合物烷基化的可能性。
