title: Region and amino acid residues required for Rad51C binding in the human Xrcc3 protein.
abstract: The Xrcc3 protein, which is required for the homologous recombinational repair of damaged DNA, forms a complex with the Rad51C protein in human cells. Mutations in either the Xrcc3 or Rad51C gene cause extreme sensitivity to DNA-damaging agents and generate the genomic instability frequently found in tumors. In the present study, we found that the Xrcc3 segment containing amino acid residues 63-346, Xrcc3(63-346), is the Rad51C-binding region. Biochemical analyses revealed that Xrcc3(63-346) forms a complex with Rad51C, and the Xrcc3(63-346)- Rad51C complex possesses ssDNA and dsDNA binding abilities comparable to those of the full-length Xrcc3-Rad51C complex. Based on the structure of RecA, which is thought to be the ancestor of Xrcc3, six Xrcc3 point mutants were designed. Two-hybrid and biochemical analyses of the Xrcc3 point mutants revealed that Tyr139 and Phe249 are essential amino acid residues for Rad51C binding. Superposition of the Xrcc3 Tyr139 and Phe249 residues on the RecA structure suggested that Tyr139 may function to ensure proper folding and Phe249 may be important to constitute the Rad51C-binding interface in Xrcc3.
authors.affiliation: RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kurumizaka
authors.forename: Rima
authors.initials: R
authors.lastname: Enomoto
authors.forename: Maki
authors.initials: M
authors.lastname: Nakada
authors.forename: Keiko
authors.initials: K
authors.lastname: Eda
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
authors.forename: Takehiko
authors.initials: T
authors.lastname: Shibata
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010649:Phenylalanine / Q000235:genetics; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D014443:Tyrosine / Q000235:genetics
chemical_list: D000596:Amino Acids; D004268:DNA-Binding Proteins; C448305:RAD51C protein, human; C110901:X-ray repair cross complementing protein 3; D014443:Tyrosine; D010649:Phenylalanine