Identification of a second DNA binding site in the human Rad52 protein
Identification of a second DNA binding site in the human Rad52 protein
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DOI：
10.1074/jbc.m802204200
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发表时间：
2008-08-29
影响因子：
4.8
通讯作者：
Yokoyama, Shigeyuki
中科院分区：
文献类型：
--
作者：
Kagawa, Wataru;Kagawa, Ako;Yokoyama, Shigeyuki
Rad 52在同源依赖性双链断裂修复中起重要作用。各种研究已经确定了Rad 52在Rad 51依赖性和Rad 51非依赖性修复过程中的功能。然而，Rad 52在这些过程中的精确分子机制仍然未知。在本研究中，我们已经确定了一个新的DNA结合位点内Rad 52的结构为基础的丙氨酸扫描诱变。该位点与先前确定的推定单链DNA结合位点紧密对齐。该位点的突变削弱了Rad 52-单链DNA复合物与双链DNA形成三元复合物并随后催化D环形成的能力。我们发现Rad 52将正超螺旋引入双链DNA中，并且第二个DNA结合位点对于这种活性是必需的。我们的研究结果表明，Rad 52对齐两个重组DNA分子内的第一和第二个DNA结合位点，以刺激同源性搜索和链侵入过程。
Rad52 plays essential roles in homology-dependent double-strand break repair. Various studies have established the functions of Rad52 in Rad51-dependent and Rad51-independent repair processes. However, the precise molecular mechanisms of Rad52 in these processes remain unknown. In the present study we have identified a novel DNA binding site within Rad52 by a structure-based alanine scan mutagenesis. This site is closely aligned with the putative single-stranded DNA binding site determined previously. Mutations in this site impaired the ability of the Rad52-single-stranded DNA complex to form a ternary complex with double-stranded DNA and subsequently catalyze the formation of D-loops. We found that Rad52 introduces positive supercoils into double-stranded DNA and that the second DNA binding site is essential for this activity. Our findings suggest that Rad52 aligns two recombining DNA molecules within the first and second DNA binding sites to stimulate the homology search and strand invasion processes.