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Identification of fidelity-governing factors in human recombinases DMC1 and RAD51 from cryo-EM structures
Nature Communications ( IF 16.6 ) Pub Date : 2021-01-14 , DOI: 10.1038/s41467-020-20258-1
Shih-Chi Luo , Hsin-Yi Yeh , Wei-Hsuan Lan , Yi-Min Wu , Cheng-Han Yang , Hao-Yen Chang , Guan-Chin Su , Chia-Yi Lee , Wen-Jin Wu , Hung-Wen Li , Meng-Chiao Ho , Peter Chi , Ming-Daw Tsai

Both high-fidelity and mismatch-tolerant recombination, catalyzed by RAD51 and DMC1 recombinases, respectively, are indispensable for genomic integrity. Here, we use cryo-EM, MD simulation and functional analysis to elucidate the structural basis for the mismatch tolerance of DMC1. Structural analysis of DMC1 presynaptic and postsynaptic complexes suggested that the lineage-specific Loop 1 Gln244 (Met243 in RAD51) may help stabilize DNA backbone, whereas Loop 2 Pro274 and Gly275 (Val273/Asp274 in RAD51) may provide an open “triplet gate” for mismatch tolerance. In support, DMC1-Q244M displayed marked increase in DNA dynamics, leading to unobservable DNA map. MD simulation showed highly dispersive mismatched DNA ensemble in RAD51 but well-converged DNA in DMC1 and RAD51-V273P/D274G. Replacing Loop 1 or Loop 2 residues in DMC1 with RAD51 counterparts enhanced DMC1 fidelity, while reciprocal mutations in RAD51 attenuated its fidelity. Our results show that three Loop 1/Loop 2 residues jointly enact contrasting fidelities of DNA recombinases.



中文翻译:

从冷冻EM结构鉴定人重组酶DMC1和RAD51中的保真因子

分别由RAD51和DMC1重组酶催化的高保真和错配耐受重组对于基因组完整性而言都是必不可少的。在这里,我们使用cryo-EM,MD仿真和功能分析来阐明DMC1不匹配容忍的结构基础。DMC1突触前和突触后复合物的结构分析表明,谱系特异性Loop 1 Gln244(RAD51中的Met243)可能有助于稳定DNA骨架,而Loop 2 Pro274和Gly275(RAD51中的Val273 / Asp274)可能为不匹配容差。在支持下,DMC1-Q244M显示出明显的DNA动力学增强,导致无法观察到DNA图。MD模拟显示RAD51中高度分散的错配DNA集合,而DMC1和RAD51-V273P / D274G中的DNA收敛良好。用RAD51对应物替换DMC1中的Loop 1或Loop 2残基可增强DMC1的保真度,而RAD51中的相互突变会减弱其保真度。我们的结果表明,三个Loop 1 / Loop 2残基共同构成了DNA重组酶的对比保真度。

更新日期:2021-01-14
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