Real-time tracking reveals catalytic roles for the two DNA binding sites of Rad51
Real-time tracking reveals catalytic roles for the two DNA binding sites of Rad51
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DOI：
10.1038/s41467-020-16750-3
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发表时间：
2020-06-11
影响因子：
16.6
通讯作者：
Iwasaki, Hiroshi
中科院分区：
文献类型：
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作者：
Ito, Kentaro;Murayama, Yasuto;Iwasaki, Hiroshi
在同源重组过程中，Rad51在单链DNA上形成核蛋白丝，促进DNA链交换。该丝结合双链DNA (dsDNA)，寻找同源性，并促进互补链的转移，产生新的异双链。链交换通过两个不同的三链中间体C1和C2进行。C1含有完整的供体双链DNA，而C2含有新形成的异双链DNA。在这里，我们发现保守的DNA结合基序，即Rad51位点I上的环1 （L1）和环2 （L2）在这一过程中发挥了不同的作用。L1参与C1复合物的形成，而L2介导C1- c2的转变，产生异双工。另一个DNA结合基序，位点II，作为初始Rad51细丝形成和供体dsDNA结合的DNA进入位置。我们的研究为真核reca家族重组酶介导的链交换催化过程提供了一个全面的分子模型。Rad51驱动DNA链交换，这是重组DNA修复的中心反应。Rad51的两个位点负责DNA结合，但这些位点的功能已被证明是难以捉摸的。在这里，作者采用实时分析来揭示Rad51的两个DNA结合位点的催化作用。
During homologous recombination, Rad51 forms a nucleoprotein filament on single-stranded DNA to promote DNA strand exchange. This filament binds to double-stranded DNA (dsDNA), searches for homology, and promotes transfer of the complementary strand, producing a new heteroduplex. Strand exchange proceeds via two distinct three-strand intermediates, C1 and C2. C1 contains the intact donor dsDNA whereas C2 contains newly formed heteroduplex DNA. Here, we show that the conserved DNA binding motifs, loop 1 (L1) and loop 2 (L2) in site I of Rad51, play distinct roles in this process. L1 is involved in formation of the C1 complex whereas L2 mediates the C1-C2 transition, producing the heteroduplex. Another DNA binding motif, site II, serves as the DNA entry position for initial Rad51 filament formation, as well as for donor dsDNA incorporation. Our study provides a comprehensive molecular model for the catalytic process of strand exchange mediated by eukaryotic RecA-family recombinases. Rad51 drives DNA strand exchange, the central reaction in recombinational DNA repair. Two sites of Rad51 are responsible for DNA binding, but the function of these sites has proven elusive. Here, the authors employ real-time assays to reveal catalytic roles for the two DNA binding sites of Rad51.