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Identification of fidelity-governing factors in human recombinases DMC1 and RAD51 from cryo-EM structures

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 Homail, Peter Chimail & Ming-Daw Tsaimail

 

Nature Communications, 12, Article number: 115 (2021)

https://www.nature.com/articles/s41467-020-20258-1

 

Abstract
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.