Functional domains of E.coli RecA protein for D-loop formation.

大肠杆菌RecA蛋白的体外反应研究已成为了解同源DNA重组分子机制的前沿研究方向。在上述课题资助的研究中，我们将重点放在了RecA蛋白的分子结构分析上，以阐明RecA蛋白在链转移反应中的分子机制。在1988和1989财年，我们获得了如下结果：1)通过分析位点定向诱变产生的几种phi80ci抑制蛋白的改变，发现RecA蛋白不参与抑制蛋白水解的催化机制。RecA蛋白通过改变其构象来增强抑制因子的自裂活性，这表明RecA蛋白具有蛋白酶的任何功能结构域。2)从c端缺失25个氨基酸残基的RecA5327蛋白与单链和双链DNA结合的亲和力高于野生型RecA蛋白。其他证据支持c端区负调控RecA蛋白的DNA结合活性。3)地区从40。60。富含疏水氨基酸残基的n端残基在RecA-RecA相互作用中起作用。基于这些结果，我们构建了RecA蛋白的头到头二聚体模型，该模型假设RecA蛋白包含一个DNA结合结构域，两个RecA-RecA相互作用结构域，以及用于解释RecA蛋白形成d环的分子机制的atp酶结构域。

The study of in vitro reaction of the RecA protein of Escherichia coli has been playing a frontier role for understanding the molecular mechanism of homologous DNA recombination. In the study funded under the title described above, we focused our efforts on the analyses of molecular structure of the RecA protein to elucidate the molecular mechanism of RecA protein in the strand transfer reaction. In the fiscal years of 1988 and 1989, we obtained the results as described below. 1) Through the analyses of several altered phi8O cI repressor proteins generated by the site directed mutagenesis, it was revealed that the RecA protein does not involved in the catalytic mechanism of the proteolysis of the repressor. The RecA protein enhances the autocleavage activity of the repressor by changing its conformation, suggesting that the RecA protein has any functional donmains for protease. 2) The RecA5327 protein which is deleted 25 amino acids residues from its C-terminal bound to both of single and double stranded DNA with higher affinities than the wild-type RecA protein. Other evidences supporting that the C-terminal region negatively regulates the DNA binding activity of RecA protein were obtained. 3) The region from 40th. to 60th. residues from N-terminal where is rich in hydrophobic amino acid residues functions for RecA-RecA interaction. From these results, we constructed the Head to Head Dimer Model of RecA protein, in which the RecA protein is postulated to contain one DNA binding domain, two domains for RecA-RecA interactions, ATPase domain for expaining the molecular mechanism of RecA protein in forming D-loop.