title: Functional analyses of the C-terminal half of the Saccharomyces cerevisiae Rad52 protein.
abstract: The Saccharomyces cerevisiae Rad52 protein is essential for efficient homologous recombination (HR). An important role of Rad52 in HR is the loading of Rad51 onto replication protein A-coated single-stranded DNA (ssDNA), which is referred to as the recombination mediator activity. In vitro, Rad52 displays additional activities, including self-association, DNA binding and ssDNA annealing. Although Rad52 has been a subject of extensive genetic, biochemical and structural studies, the mechanisms by which these activities are coordinated in the various roles of Rad52 in HR remain largely unknown. In the present study, we found that an isolated C-terminal half of Rad52 disrupted the Rad51 oligomer and formed a heterodimeric complex with Rad51. The Rad52 fragment inhibited the binding of Rad51 to double-stranded DNA, but not to ssDNA. The phenylalanine-349 and tyrosine-409 residues present in the C-terminal half of Rad52 were critical for the interaction with Rad51, the disruption of Rad51 oligomers, the mediator activity of the full-length protein and for DNA repair in vivo in the presence of methyl methanesulfonate. Our studies suggested that phenylalanine-349 and tyrosine-409 are key residues in the C-terminal half of Rad52 and probably play an important role in the mediator activity.
authors.affiliation: Department of Interdisciplinary Science and Engineering, Program in Chemistry and Life Science, School of Science and Engineering, Meisei University, 2-1-1 Hodokubo, Hino-shi, Tokyo 191-8506, Japan, Department of Applied Biological Science, Nihon University College of Bioresource Sciences, Fujisawa-shi, Kanagawa 252-0880, Japan, Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan and Cellular and Molecular Biology Laboratory, RIKEN, Wako-shi, Saitama 351-0198, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Kagawa
authors.forename: Naoto
authors.initials: N
authors.lastname: Arai
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Ichikawa
authors.forename: Kengo
authors.initials: K
authors.lastname: Saito
authors.forename: Shusei
authors.initials: S
authors.lastname: Sugiyama
authors.forename: Mika
authors.initials: M
authors.lastname: Saotome
authors.forename: Takehiko
authors.initials: T
authors.lastname: Shibata
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kurumizaka
mesh_terms: D000595:Amino Acid Sequence; D004247:DNA / Q000378:metabolism; D004260:DNA Repair; D059765:Homologous Recombination; D008969:Molecular Sequence Data; D009154:Mutation; D010649:Phenylalanine / Q000235:genetics; D055503:Protein Multimerization; D051135:Rad51 Recombinase / Q000378:metabolism; D051721:Rad52 DNA Repair and Recombination Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014443:Tyrosine / Q000235:genetics
chemical_list: C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D014443:Tyrosine; D010649:Phenylalanine; D004247:DNA; D051135:Rad51 Recombinase