Department of Chemistry and Biochemistry, University of Delaware, Newark, DE 19716, USA.
J Mol Biol. 2011 Jan 7;405(1):77-91. doi: 10.1016/j.jmb.2010.10.042. Epub 2010 Nov 5.
Rad51 and Rad54 play crucial roles during homologous recombination. The biochemical activities of human Rad51 (hRad51) and human Rad54 (hRad54) and their interactions with each other are well documented. However, it is not known how these two proteins work together to bypass heterologous sequences; i.e. mismatched base pairs, during homologous recombination. In this study, we used a fluorescence resonance energy transfer assay to monitor homologous recombination processes in real time so that the interactions between hRad54 and hRad51 during DNA strand exchange and branch migration, which are two core steps of homologous recombination, could be characterized. Our results indicate that hRad54 can facilitate hRad51-promoted strand exchange through various degrees of mismatching. We propose that the main roles of hRad51 in homologous recombination is to initiate the homology recognition and strand-exchange steps and those of hRad54 are to promote efficient branch migration, bypass potential mismatches and facilitate long-range strand exchanges through branch migration of Holliday junctions.
Rad51 和 Rad54 在同源重组中发挥着至关重要的作用。人类 Rad51（hRad51）和人类 Rad54（hRad54）的生化活性及其相互作用已得到充分证实。然而，目前尚不清楚这两种蛋白如何协同作用以在同源重组过程中绕过异源序列（即错配碱基对）。在这项研究中，我们使用荧光共振能量转移测定法实时监测同源重组过程，从而能够对 DNA 链交换和分支迁移这两个同源重组的核心步骤中 hRad54 和 hRad51 之间的相互作用进行特征描述。研究结果表明，hRad54 可以通过不同程度的错配促进 hRad51 介导的链交换。我们提出，hRad51 在同源重组中的主要作用是启动同源性识别和链交换步骤，而 hRad54 的主要作用是促进有效的分支迁移，绕过潜在的错配并通过 Holliday 连接点的分支迁移促进长距离链交换。
