Hatahet Z, Kow Y W, Purmal A A, Cunningham R P, Wallace S S
Department of Microbiology and Molecular Genetics, University of Vermont, Burlington 05405.
J Biol Chem. 1994 Jul 22;269(29):18814-20.
5-Hydroxy-2'-deoxycytidine (5-OHdC) and 5-hydroxy-2'-deoxyuridine (5-OHdU) are major products of oxidative DNA damage with mutagenic potential. Until now, no enzymatic activity responsible for their removal has been identified. We report here that both 5-OHdC and 5-OHdU are substrates for Escherichia coli endonuclease III and formamidopyrimidine DNA N-glycosylase (FPG). 5-OHdU is also a substrate for uracil DNA N-glycosylase. Consistent with their mechanisms of action on previously described substrates, endonuclease III removes 5-OHdC and 5-OHdU via a N-glycosylase/beta-elimination reaction, FPG follows a N-glycosylase/beta,delta-elimination reaction, and uracil N-glycosylase removes 5-OHdU by N-glycosylase action leaving behind an abasic site. Endonuclease III removes both lesions more efficiently than FPG, and both endonuclease III and FPG remove 5-OHdC slightly more efficiently than 5-OHdU. Uracil DNA N-glycosylase removes 5-OHdU more efficiently than the other two enzymes and has no activity on 5-OHdC even when present in great excess. Analysis of crude extracts obtained from wild type and endonuclease III deletion mutants of E. coli correlated well with data obtained with the purified enzymes.
5-羟基-2'-脱氧胞苷（5-OHdC）和5-羟基-2'-脱氧尿苷（5-OHdU）是具有诱变潜力的氧化性DNA损伤的主要产物。到目前为止，尚未鉴定出负责去除它们的酶活性。我们在此报告，5-OHdC和5-OHdU都是大肠杆菌内切核酸酶III和甲酰胺嘧啶DNA N-糖基化酶（FPG）的底物。5-OHdU也是尿嘧啶DNA N-糖基化酶的底物。与它们对先前描述的底物的作用机制一致，内切核酸酶III通过N-糖基化酶/β-消除反应去除5-OHdC和5-OHdU，FPG遵循N-糖基化酶/β,δ-消除反应，尿嘧啶N-糖基化酶通过N-糖基化酶作用去除5-OHdU，留下一个无碱基位点。内切核酸酶III比FPG更有效地去除这两种损伤，并且内切核酸酶III和FPG去除5-OHdC的效率略高于5-OHdU。尿嘧啶DNA N-糖基化酶比其他两种酶更有效地去除5-OHdU，并且即使大量存在时对5-OHdC也没有活性。对从大肠杆菌野生型和内切核酸酶III缺失突变体获得的粗提物的分析与用纯化酶获得的数据相关性良好。
