Moe Ane, Ringvoll Jeanette, Nordstrand Line M, Eide Lars, Bjørås Magnar, Seeberg Erling, Rognes Torbjørn, Klungland Arne
Centre for Molecular Biology and Neuroscience and Institute of Medical Microbiology, University of Oslo, Rikshospitalet, N-0027 Oslo, Norway.
Nucleic Acids Res. 2003 Jul 15;31(14):3893-900. doi: 10.1093/nar/gkg472.
Deamination of DNA bases can occur spontaneously, generating highly mutagenic lesions such as uracil and hypoxanthine. In Escherichia coli two enzymes initiate repair at hypoxanthine residues in DNA. The alkylbase DNA glycosylase, AlkA, initiates repair by removal of the damaged base, whereas endonuclease V, Endo V, hydrolyses the second phosphodiester bond 3' to the lesion. We have identified and characterised a mouse cDNA with striking homology to the E.coli nfi gene, which also has significant similarities to motifs required for catalytic activity of the UvrC endonuclease. The 37-kDa mouse enzyme (mEndo V) incises the DNA strand at the second phosphodiester bond 3' to hypoxanthine- and uracil-containing nucleotides. The activity of mEndo V is elevated on single-stranded DNA substrate in vitro. Expression of the mouse protein in a DNA repair-deficient E.coli alkA nfi strain suppresses its spontaneous mutator phenotype. We suggest that mEndo V initiates an alternative excision repair pathway for hypoxanthine removal. It thus appears that mEndo V has properties overlapping the function of alkylbase DNA glycosylase (Aag) in repair of deaminated adenine, which to some extent could explain the absence of phenotypic abnormalities associated with Aag knockout in mice.
DNA碱基的脱氨基作用可自发发生，产生高度诱变的损伤，如尿嘧啶和次黄嘌呤。在大肠杆菌中，有两种酶启动对DNA中次黄嘌呤残基的修复。烷基碱基DNA糖基化酶AlkA通过去除受损碱基启动修复，而核酸内切酶V（Endo V）则水解损伤位点3'端的第二个磷酸二酯键。我们已经鉴定并表征了一个与大肠杆菌nfi基因具有显著同源性的小鼠cDNA，该基因与UvrC核酸内切酶催化活性所需的基序也有显著相似性。这种37 kDa的小鼠酶（mEndo V）在含次黄嘌呤和尿嘧啶的核苷酸的3'端第二个磷酸二酯键处切割DNA链。在体外，mEndo V在单链DNA底物上的活性会升高。在DNA修复缺陷的大肠杆菌alkA nfi菌株中表达小鼠蛋白可抑制其自发突变体表型。我们认为mEndo V启动了一条用于去除次黄嘌呤的替代切除修复途径。因此，mEndo V的特性似乎与烷基碱基DNA糖基化酶（Aag）在脱氨基腺嘌呤修复中的功能重叠，这在一定程度上可以解释小鼠中与Aag基因敲除相关的表型异常的缺失。
