Human Rad54 protein stimulates DNA strand exchange activity of hRad51 protein in the presence of Ca2+
Human Rad54 protein stimulates DNA strand exchange activity of hRad51 protein in the presence of Ca2+
复制标题
DOI：
10.1074/jbc.m410244200
复制
发表时间：
2004-12-10
影响因子：
4.8
通讯作者：
Mazin, AV
中科院分区：
文献类型：
--
作者：
Mazina, OM;Mazin, AV
RAD51和RAD54蛋白在真核生物的同源重组中起着关键作用。最近，我们报道了在体外，人类RAD51蛋白需要钙离子来形成一个活性的核蛋白细丝，这对于寻找同源DNA和DNA链交换是重要的，这是同源重组的两个关键步骤。我们发现，hRad54蛋白也需要钙离子才能有效地刺激hRad51蛋白的DNA链交换活性。这一发现表明，在体外，Ca~(2+)是哺乳动物同源重组蛋白促进DNA链交换的普遍辅因子。我们进一步研究了依赖hRad54的DNA链交换的刺激作用。刺激机制可能包括hRad54蛋白与hRad51核蛋白细丝的特异性相互作用。我们的结果表明，hRad54蛋白显著刺激dsDNA与细丝的同源非同源共聚集，这代表了寻找同源DNA的关键步骤。结果表明，hRad54蛋白是hRad51-ssDNA细丝的dsDNA通道，在寻找同源序列的过程中促进dsDNA的结合和依赖于ATP水解酶的转位。
Rad51 and Rad54 proteins play a key role in homologous recombination in eukaryotes. Recently, we reported that Ca2+ is required in vitro for human Rad51 protein to form an active nucleoprotein filament that is important for the search of homologous DNA and for DNA strand exchange, two critical steps of homologous recombination. Here we find that Ca2+ is also required for hRad54 protein to effectively stimulate DNA strand exchange activity of hRad51 protein. This finding identifies Ca2+ as a universal cofactor of DNA strand exchange promoted by mammalian homologous recombination proteins in vitro. We further investigated the hRad54-dependent stimulation of DNA strand exchange. The mechanism of stimulation appeared to include specific interaction of hRad54 protein with the hRad51 nucleoprotein filament. Our results show that hRad54 protein significantly stimulates homology-independent coaggregation of dsDNA with the filament, which represents an essential step of the search for homologous DNA. The results obtained indicate that hRad54 protein serves as a dsDNA gateway for the hRad51-ssDNA filament, promoting binding and an ATP hydrolysis-dependent translocation of dsDNA during the search for homologous sequences.