title: Physical interaction between replication protein A and Rad51 promotes exchange on single-stranded DNA.
abstract: Replication protein A (RPA) is displaced from single-stranded DNA (ssDNA) by Rad51 during the initiation of homologous recombination. Interactions between these proteins have been reported, but the functional significance of the direct RPA-Rad51 interaction has yet to be elucidated. We have identified and characterized the interaction between DNA-binding domain A of RPA (RPA70A) and the N-terminal domain of Rad51 (Rad51N). NMR chemical shift mapping showed that Rad51N binds to the ssDNA-binding site of RPA70A, suggesting a competitive mechanism for the displacement of RPA from ssDNA by Rad51. A structure of the RPA70A-Rad51N complex was generated by experimentally guided modeling and then used to design mutations that disrupt the binding interface. Functional ATP hydrolysis assays were performed for wild-type Rad51 and a mutant defective in binding RPA. Rates of RPA displacement for the mutant were significantly below those of wild-type Rad51, suggesting that a direct RPA-Rad51 interaction is involved in displacing RPA in the initiation stage of genetic recombination.
authors.affiliation: Departments of Biochemistry and Physics and the Center for Structural Biology, Vanderbilt University, Nashville, Tennessee 37232-8725, USA.
authors.forename: Melissa E
authors.initials: ME
authors.lastname: Stauffer
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Chazin
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D017354:Point Mutation; D051135:Rad51 Recombinase; D011995:Recombination, Genetic; D051716:Replication Protein A
chemical_list: D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D051716:Replication Protein A; D000255:Adenosine Triphosphate; D051135:Rad51 Recombinase