Biochemical analysis of the N-terminal domain of human RAD54B.
The human RAD54B protein is a paralog of the RAD54 protein, which plays important roles in homologous recombination. RAD54B contains an N-terminal region outside the SWI2/SNF2 domain that shares less conservation with the corresponding region in RAD54. The biochemical roles of this region of RAD54B are not known, although the corresponding region in RAD54 is known to physically interact with RAD51. In the present study, we have biochemically characterized an N-terminal fragment of RAD54B, consisting of amino acid residues 26-225 (RAD54B(26-225)). This fragment formed a stable dimer in solution and bound to branched DNA structures. RAD54B(26-225) also interacted with DMC1 in both the presence and absence of DNA. Ten DMC1 segments spanning the entire region of the DMC1 sequence were prepared, and two segments, containing amino acid residues 153-214 and 296-340, were found to directly bind to the N-terminal domain of RAD54B. A structural alignment of DMC1 with the Methanococcus voltae RadA protein, a homolog of DMC1 in the helical filament form, indicated that these RAD54B-binding sites are located near the ATP-binding site at the monomer-monomer interface in the DMC1 helical filament. Thus, RAD54B binding may affect the quaternary structure of DMC1. These observations suggest that the N-terminal domain of RAD54B plays multiple roles of in homologous recombination.
Sarai
Naoyuki
N
Systems and Structural Biology Center, Yokohama Institute, RIKEN, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.
Kagawa
Wataru
W
Fujikawa
Norie
N
Saito
Kengo
K
Hikiba
Juri
J
Tanaka
Kozo
K
Miyagawa
Kiyoshi
K
Kurumizaka
Hitoshi
H
Yokoyama
Shigeyuki
S
D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry* / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D051135:Rad51 Recombinase / Q000378:metabolism
D018797:Cell Cycle Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C491221:RAD54B protein, human; D004247:DNA; D051135:Rad51 Recombinase; C074556:DMC1 protein, human; D004265:DNA Helicases