Hang B, Chenna A, Rao S, Singer B
Life Science Division, Donner Laboratory, Lawrence Berkeley National Laboratory, University of California, Berkeley 94720, USA.
Carcinogenesis. 1996 Jan;17(1):155-7. doi: 10.1093/carcin/17.1.155.
We previously reported our finding that human cells contain glycosylase activity toward all four etheno bases formed in DNA by chloroacetaldehyde and related bi-functional aldehydes. By enzyme purification, including FPLC, we isolated two separate glycosylase activities for 1,N6-ethenoadenine (epsilon A) and for 3,N4-ethenocytosine (epsilon C) respectively, from crude HeLa cell-free extracts, which also contained a number of well-described glycosylases. When Mono-S FPLC purified proteins were assayed against defined oligomers containing either epsilon A or epsilon C, it was found that epsilon A and epsilon C glycosylases were completely separated. It could also be demonstrated that each enzyme bound to and cut only epsilon A- or epsilon C-containing oligomers respectively. There was no overlap in specificity for these two substrates. Several other human glycosylase substrates were also tested and none were cleaved by epsilon C glycosylase. The epsilon C glycosylase activity identified in the present study apparently represents a previously unknown glycosylase. This work also suggests that enzyme recognition of closely related DNA adducts may depend upon subtle changes in local conformation.
我们之前报道了一项发现，即人类细胞对氯乙醛及相关双功能醛在DNA中形成的所有四种乙烯基碱基具有糖基化酶活性。通过包括快速蛋白质液相色谱（FPLC）在内的酶纯化方法，我们从HeLa细胞无细胞粗提物中分别分离出了针对1,N6-乙烯基腺嘌呤（εA）和3,N4-乙烯基胞嘧啶（εC）的两种独立的糖基化酶活性，该粗提物中还含有多种已被充分描述的糖基化酶。当用含有εA或εC的特定寡聚物对Mono-S FPLC纯化的蛋白质进行检测时，发现εA和εC糖基化酶被完全分离。还能证明每种酶分别仅与含有εA或εC的寡聚物结合并切割，这两种底物在特异性上没有重叠。还测试了其他几种人类糖基化酶底物，没有一种被εC糖基化酶切割。本研究中鉴定出的εC糖基化酶活性显然代表了一种此前未知的糖基化酶。这项工作还表明，酶对密切相关的DNA加合物的识别可能取决于局部构象的细微变化。
