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Mechanism of homologous recombination from the RecA–ssDNA/dsDNA structures

Abstract

The RecA family of ATPases mediates homologous recombination, a reaction essential for maintaining genomic integrity and for generating genetic diversity. RecA, ATP and single-stranded DNA (ssDNA) form a helical filament that binds to double-stranded DNA (dsDNA), searches for homology, and then catalyses the exchange of the complementary strand, producing a new heteroduplex. Here we have solved the crystal structures of the Escherichia coli RecA–ssDNA and RecA–heteroduplex filaments. They show that ssDNA and ATP bind to RecA–RecA interfaces cooperatively, explaining the ATP dependency of DNA binding. The ATP γ-phosphate is sensed across the RecA–RecA interface by two lysine residues that also stimulate ATP hydrolysis, providing a mechanism for DNA release. The DNA is underwound and stretched globally, but locally it adopts a B-DNA-like conformation that restricts the homology search to Watson–Crick-type base pairing. The complementary strand interacts primarily through base pairing, making heteroduplex formation strictly dependent on complementarity. The underwound, stretched filament conformation probably evolved to destabilize the donor duplex, freeing the complementary strand for homology sampling.

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Figure 1: Structure of the presynaptic nucleoprotein filament.
Figure 2: Each nucleotide triplet is bound by three consecutive RecA protomers.
Figure 3: The non-hydrolysable ATP analogue ADP-AlF 4 binds at a RecA–RecA interface.
Figure 4: Structure of the postsynaptic nucleoprotein filament.
Figure 5: Complementary-strand binding.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The coordinates and structure factors have been deposited in the Protein Data Bank under accession codes 3CMW (RecA5–(dT)15), 3CMU (RecA6–(dT)18), 3CMX (RecA5–(dT)15–(dA)12), 3CMT (RecA5–d(T5C3AC2T4)–d(G2TG3)) and 3CMV (RecA4).

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Acknowledgements

We thank D. King for mass spectroscopic analysis; H. Erdjument-Bromage for N-terminal sequencing; the staff of the Advanced Photon Source ID24 beamlines for help with data collection; M. Minto for administrative assistance; and the members of the Pavletich laboratory for help and discussions. This work was supported by the NIH and the Howard Hughes Medical Institute.

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Correspondence to Nikola P. Pavletich.

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The file contains Supplementary Tables 1-5, Supplementary Figures 1-11 with Legends and additional references (PDF 2798 kb)

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Chen, Z., Yang, H. & Pavletich, N. Mechanism of homologous recombination from the RecA–ssDNA/dsDNA structures. Nature 453, 489–494 (2008). https://doi.org/10.1038/nature06971

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