Novel repair activities of AlkA (3-methyladenine DNA glycosylase II) and endonuclease VIII for xanthine and oxanine, guanine lesions induced by nitric oxide and nitrous acid.
Novel repair activities of AlkA (3-methyladenine DNA glycosylase II) and endonuclease VIII for xanthine and oxanine, guanine lesions induced by nitric oxide and nitrous acid.
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DOI：
10.1093/nar/gkf630
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发表时间：
2002-11
影响因子：
14.9
通讯作者：
H. Terato;A. Masaoka;K. Asagoshi;A. Honsho;Y. Ohyama;Toshinori Suzuki;M. Yamada;K. Makino;Kazuo Yamamoto;H. Ide
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文献类型：
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作者：
H. Terato;A. Masaoka;K. Asagoshi;A. Honsho;Y. Ohyama;Toshinori Suzuki;M. Yamada;K. Makino;Kazuo Yamamoto;H. Ide
DNA 中的鸟嘌呤被一氧化氮 (NO) 和亚硝酸等氮氧化物亚硝化，导致形成黄嘌呤 (Xan) 和恶嘌呤 (Oxa)，可能具有细胞毒性和致突变性病变。在本研究中，我们检测了大肠杆菌 DNA N-糖基化酶对 Xan 和 Oxa 的修复能力。使用含有 Xan 和 Oxa 的确定底物进行的切口测定表明，AlkA [与核酸内切酶 (Endo) IV 组合]和 Endo VIII 识别测试酶中的 Xan。 AlkA 对 Xan 的活性 (V(max)/K(m)) 比对 7-甲基鸟嘌呤的活性低 5 倍，而 Endo VIII 的活性比对胸腺嘧啶二醇低 50 倍。 AlkA 和 Endo VIII 对 Xan 的活性通过从底物中释放 [(3)H]Xan 得到进一步证实。用 N-甲基-N'-硝基-N-亚硝基胍处理大肠杆菌可增加 alkA(+) 菌株细胞提取物中的 Xan 切除活性，但不会增加 alkA(-) 菌株的细胞提取物中的 Xan 切除活性。相对于野生型菌株，alkA 和 nei（Endo VIII 基因）双突变体（而非单突变体）表现出对亚硝酸的敏感性增加。 AlkA 和 Endo VIII 也表现出对 Oxa 的切除活性，但该活性远低于 Xan。
Nitrosation of guanine in DNA by nitrogen oxides such as nitric oxide (NO) and nitrous acid leads to formation of xanthine (Xan) and oxanine (Oxa), potentially cytotoxic and mutagenic lesions. In the present study, we have examined the repair capacity of DNA N-glycosylases from Escherichia coli for Xan and Oxa. The nicking assay with the defined substrates containing Xan and Oxa revealed that AlkA [in combination with endonuclease (Endo) IV] and Endo VIII recognized Xan in the tested enzymes. The activity (V(max)/K(m)) of AlkA for Xan was 5-fold lower than that for 7-methylguanine, and that of Endo VIII was 50-fold lower than that for thymine glycol. The activity of AlkA and Endo VIII for Xan was further substantiated by the release of [(3)H]Xan from the substrate. The treatment of E.coli with N-methyl-N'-nitro-N-nitrosoguanidine increased the Xan-excising activity in the cell extract from alkA(+) but not alkA(-) strains. The alkA and nei (the Endo VIII gene) double mutant, but not the single mutants, exhibited increased sensitivity to nitrous acid relative to the wild type strain. AlkA and Endo VIII also exhibited excision activity for Oxa, but the activity was much lower than that for Xan.