SEQUENCING, TARGETED MUTAGENESIS AND EXPRESSION OF A RECA GENE REQUIRED FOR THE EXTREME RADIORESISTANCE OF DEINOCOCCUS RADIODURANS
SEQUENCING, TARGETED MUTAGENESIS AND EXPRESSION OF A RECA GENE REQUIRED FOR THE EXTREME RADIORESISTANCE OF DEINOCOCCUS RADIODURANS
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DOI：
10.1016/0378-1119(94)90124-4
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发表时间：
1994-04-08
期刊：
影响因子：
3.5
通讯作者：
MINTON, KW
中科院分区：
文献类型：
--
作者：
GUTMAN, PD;CARROLL, JD;MINTON, KW
耐辐射异常球菌和同一属的其他成员对电离辐射和许多其他破坏DNA的物质具有极强的抵抗力。发现通过化学诱变产生的DNA损伤敏感性和天然转化缺陷型菌株（菌株rec 30）在与大肠杆菌的recA具有延伸同源性的基因中存在缺陷。在用含有来自野生型（wt）D的该异常球菌recA基因的染色体DNA片段转化后，在rec 30突变体中恢复了耐辐射菌的DNA损伤抗性和完全转化能力。使用异常球菌recA基因的靶向插入诱变来构建与wt.插入突变体是从菌株rec 30表型上无法区分的，这表明recA缺陷单独负责观察到的表型改变。例如，在电离辐射的情况下，wt的D-37约为1.75 Mrad，而rec 30和插入突变体的D-37均为25 krad，减少了70倍。有证据表明，异常球菌recA基因在E.大肠杆菌的RecA是致命的，这表明异常球菌RecA蛋白与核酸或其他细胞蛋白的相互作用模式至少部分不同于大肠杆菌的RecA。杆菌
Deinococcus radiodurans and other members of the same genus share extreme resistance to ionizing radiation and many other agents that damage DNA. A DNA damage-sensitive and natural transformation-deficient strain generated by chemical mutagenesis (strain rec30) was found to be defective in a gene that has extended homology with recA of Escherichia coli. Upon transformation with a chromosomal DNA fragment that contained this deinococcal recA gene from wild-type (wt) D. radiodurans both DNA damage resistance and full transformation competence were restored in the rec30 mutant. Targeted insertional mutagenesis of the deinococcal recA gene was used to construct a mutant isogenic with the wt. The insertional mutant was phenotypically indistinguishable from strain rec30, indicating that the recA defect alone was responsible for observed phenotypic alterations. For example, in the case of ionizing radiation, the D-37 of the wt was about 1.75 Mrad, while the D-37 of rec30 and the insertional mutant were both 25 krad, a 70-fold decrease. Evidence is presented that expression of the deinococcal recA gene in E. coli is lethal, suggesting that the mode of interaction of the deinococcal RecA protein with nucleic acids or other cellular proteins differs at least in part from RecA of E. coli.