Holliday junction-binding activity of human SPF45.
SPF45 is considered to be a bifunctional protein that functions in splicing and DNA repair. A previous genetic study reported that Drosophila SPF45 participates in the DNA-repair pathway with a RAD51-family protein, RAD201, suggesting that SPF45 may function in DNA repair by the homologous-recombination pathway. To study the function of SPF45 in homologous recombination, we purified human SPF45 and found that it preferentially binds to the Holliday junction, which is a key DNA intermediate in the homologous-recombination pathway. Deletion analyses revealed that the RNA recognition motif, which is located in the C-terminal region of human SPF45, is not involved in DNA binding. On the other hand, alanine-scanning mutagenesis identified the N-terminal lysine residues, which may be involved in Holliday junction binding by human SPF45. We also found that human SPF45 significantly binds to a RAD51 paralog, RAD51B, although it also binds to RAD51 and DMC1 with lower affinity. These biochemical results support the idea that human SPF45 functions in DNA repair by homologous recombination.
Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
Naoki
N
Horikoshi
Yuichi
Y
Morozumi
Motoki
M
Takaku
Yoshimasa
Y
Takizawa
Hitoshi
H
Kurumizaka
D018797:Cell Cycle Proteins; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D045566:DNA, Cruciform / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins; D018390:Gene Targeting; D006801:Humans; D016296:Mutagenesis; D012326:RNA Splicing; D051135:Rad51 Recombinase / Q000235:genetics* / Q000378:metabolism*; D011995:Recombination, Genetic
D018797:Cell Cycle Proteins; D045566:DNA, Cruciform; D004268:DNA-Binding Proteins; D004247:DNA; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; C074556:DMC1 protein, human