Biochemical events essential to the recombination activity of Escherichia coli RecA protein. I. Properties of the mutant RecA142 protein.
Biochemical events essential to the recombination activity of Escherichia coli RecA protein. I. Properties of the mutant RecA142 protein.
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对大肠杆菌 RecA 蛋白重组活性至关重要的生化事件。
DOI：
10.1016/0022-2836(89)90239-8
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发表时间：
1989
影响因子：
5.6
通讯作者：
Krupp,RA
中科院分区：
文献类型：
--
作者：
Kowalczykowski,SC;Burk,DL;Krupp,RA
我们对大肠杆菌RecA 142蛋白的生化特性进行了研究，该蛋白是表型上遗传重组缺陷的arecA等位基因的产物，在体外，该突变体RecA蛋白完全不能形成异源双链DNA。尽管有这个缺陷，RecA 142蛋白并不缺乏所有其他生物化学活性。RecA 142蛋白精通单链（ss）DNA结合能力、ssDNA依赖性ATP酶活性和无DNA自缔合（尽管前2个性质显示出比野生型蛋白对NaCl浓度更大的敏感性）。然而，RecA 142蛋白缺乏四个特性：（1）其ssDNA依赖性ATP酶活性完全被ssDNA结合（SSB）蛋白抑制，表明RecA 142蛋白无法与SSB蛋白有效竞争ssDNA结合位点;（2）无法促进ssDNA和双链（ds）DNA的共聚集;（3）其M13 dsDNA依赖性ATP酶活性减弱至野生型蛋白水平的约5%;（4）其不能完全形成通常由ATP诱导的高亲和力ssDNA结合状态的特征。前三个缺陷分别对应于thein vitroDNA链交换反应的突触前、突触和突触后步骤中的缺陷;第四个缺陷可能是缺陷1和3的根本基础。因此，这些性质中的一个或多个必须对体外和体内过程都很重要。
We have characterized the biochemical properties ofEscherichia coliRecA142 protein, the product of arecAallele that is phenotypically defective in genetic recombination.In vitro, this mutant RecA protein is totally defective in DNA heteroduplex formation. Despite this defect, RecA142 protein is not deficient in all other biochemical activities. RecA142 protein is proficient in single-strand (ss) DNA binding ability, ssDNA-dependent ATPase activity, and DNA-free self-association (although the first 2 properties show a greater sensitivity to NaCl concentration than does the wild-type protein). However, RecA142 protein is deficient in four properties: (1) its ssDNA-dependent ATPase activity is completely inhibited by ssDNA binding (SSB) protein, demonstrating that RecA142 protein is unable to compete effectively with SSB protein for ssDNA binding sites; (2) it is unable to promote the coaggregation of ssDNA and double-strand (ds) DNA; (3) its M13 dsDNA-dependent ATPase activity is attenuated to approximately 5% of the level of the wild-type protein; (4) it is unable fully to develop characteristics of the high-affinity ssDNA-binding state that is normally induced by ATP. The first three deficiencies correspond to defects in the presynaptic, synaptic and postsynaptic steps of thein vitroDNA strand exchange reaction, respectively; the fourth is the likely fundamental basis for defects 1 and 3. Therefore, one or more of these properties must be important to both thein vitroandin vivoprocesses.