Recombination mediator and Rad51 targeting activities of a human BRCA2 polypeptide.
Recombination mediator and Rad51 targeting activities of a human BRCA2 polypeptide.
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DOI：
10.1074/jbc.m601249200
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发表时间：
2006-04-28
期刊：
影响因子：
--
通讯作者：
Sung P
中科院分区：
文献类型：
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作者：
San Filippo J;Chi P;Sehorn MG;Etchin J;Krejci L;Sung P
BRCA 2可能通过增强损伤染色体的同源定向修复效率来发挥其肿瘤抑制功能。为了帮助确定BRCA 2的DNA修复作用，我们表达并纯化了一种多肽BRC 3/4-DBD，它含有BRC 3和BRC 4重复序列和DNA结合结构域。BRC 3/4-DBD与hRad 51相互作用，并以明显偏好单链（ss）DNA的方式结合DNA。重要的是，我们通过生物化学手段和电子显微镜证实了BRC 3/4-DBD使hRad 51在ssDNA上成核，并作为重组介体使hRad 51能够利用复制蛋白A包被的ssDNA作为重组底物。BRC 3/4-DBD的这些功能需要BRC重复序列和BRCA 2 DNA结合结构域。因此，这些结果阐明了BRCA 2在Rad 51依赖性DNA重组和修复中的作用，本文所述的实验策略对于系统地解释BRCA 2的功能应该是有价值的。
BRCA2 likely exerts its tumor suppressor function by enhancing the efficiency of the homology-directed repair of injured chromosomes. To help define the DNA repair role of BRCA2, we expressed and purified a polypeptide, BRC3/4-DBD, that harbors its BRC3 and BRC4 repeats and DNA binding domain. BRC3/4-DBD interacted with hRad51 and bound DNA with a distinct preference for single-stranded (ss) DNA. Importantly we demonstrated by biochemical means and electron microscopy that BRC3/4-DBD nucleates hRad51 onto ssDNA and acts as a recombination mediator in enabling hRad51 to utilize replication protein A-coated ssDNA as recombination substrate. These functions of BRC3/4-DBD required both the BRC repeats and the BRCA2 DNA binding domain. The results thus clarify the role of BRCA2 in Rad51-dependent DNA recombination and repair, and the experimental strategies described herein should be valuable for systematically deciphering this BRCA2 function.