Precise binding of single-stranded DNA termini by human RAD52 protein.
The human RAD52 protein, which exhibits a heptameric ring structure, has been shown to bind resected double strand breaks (DSBs), consistent with an early role in meiotic recombination and DSB repair. In this work, we show that RAD52 binds single-stranded and tailed duplex DNA molecules via precise interactions with the terminal base. When probed with hydroxyl radicals, ssDNA-RAD52 complexes exhibit a four-nucleotide repeat hypersensitivity pattern. This unique pattern is due to the interaction of RAD52 with either a 5' or a 3' terminus of the ssDNA, is sequence independent and is phased precisely from the terminal nucleotide. Hypersensitivity is observed over approximately 36 nucleotides, consistent with the length of DNA that is protected by RAD52 in nuclease protection assays. We propose that RAD52 binds DNA breaks via specific interactions with the terminal base, leading to the formation of a precisely organized ssDNA-RAD52 complex in which the DNA lies on an exposed surface of the protein. This protein-DNA arrangement may facilitate the DNA-DNA interactions necessary for RAD52-mediated annealing of complementary DNA strands.
Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Hertfordshire EN6 3LD, UK.
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Parsons
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Baumann
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Van Dyck
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West
D001665:Binding Sites; D004249:DNA Damage; D018983:DNA Footprinting; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D017665:Hydroxyl Radical; D008958:Models, Molecular; D011485:Protein Binding; D051721:Rad52 DNA Repair and Recombination Protein
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C497194:RAD52 protein, human; D051721:Rad52 DNA Repair and Recombination Protein; D017665:Hydroxyl Radical