Human RAD52 Captures and Holds DNA Strands, Increases DNA Flexibility, and Prevents Melting of Duplex DNA: Implications for DNA Recombination.
Human RAD52 promotes annealing of complementary single-stranded DNA (ssDNA). In-depth knowledge of RAD52-DNA interaction is required to understand how its activity is integrated in DNA repair processes. Here, we visualize individual fluorescent RAD52 complexes interacting with single DNA molecules. The interaction with ssDNA is rapid, static, and tight, where ssDNA appears to wrap around RAD52 complexes that promote intra-molecular bridging. With double-stranded DNA (dsDNA), interaction is slower, weaker, and often diffusive. Interestingly, force spectroscopy experiments show that RAD52 alters the mechanics dsDNA by enhancing DNA flexibility and increasing DNA contour length, suggesting intercalation. RAD52 binding changes the nature of the overstretching transition of dsDNA and prevents DNA melting, which is advantageous for strand clamping during or after annealing. DNA-bound RAD52 is efficient at capturing ssDNA in trans. Together, these effects may help key steps in DNA repair, such as second-end capture during homologous recombination or strand annealing during RAD51-independent recombination reactions.
Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, the Netherlands.
Ineke
I
Brouwer
Cancer Research Center of Marseille, CNRS UMR7258, Inserm U1068, Institut Paoli-Calmettes, Aix-Marseille Université UM105, 13273 Marseille, France.
Hongshan
H
Zhang
Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, the Netherlands.
Andrea
A
Candelli
Cancer Research Center of Marseille, CNRS UMR7258, Inserm U1068, Institut Paoli-Calmettes, Aix-Marseille Université UM105, 13273 Marseille, France.
Davide
D
Normanno
Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, the Netherlands. Electronic address: e.j.g.peterman@vu.nl.
Erwin J G
EJG
Peterman
Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, the Netherlands. Electronic address: g.j.l.wuite@vu.nl.
Gijs J L
GJL
Wuite
Cancer Research Center of Marseille, CNRS UMR7258, Inserm U1068, Institut Paoli-Calmettes, Aix-Marseille Université UM105, 13273 Marseille, France. Electronic address: mauro.modesti@inserm.fr.
Mauro
M
Modesti
D004247:DNA / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004058:Diffusion; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D009691:Nucleic Acid Denaturation; D011485:Protein Binding; D051721:Rad52 DNA Repair and Recombination Protein / Q000378:metabolism*; D011995:Recombination, Genetic
D004277:DNA, Single-Stranded; C497194:RAD52 protein, human; D051721:Rad52 DNA Repair and Recombination Protein; D049452:Green Fluorescent Proteins; D004247:DNA