Homologous Pairing and Ring and Filament Structure Formation Activities of the Human Xrcc2·Rad51D Complex*
- Hitoshi Kurumizaka‡,§,¶,
- Shukuko Ikawa‖,
- Maki Nakada‡,
- Rima Enomoto‡,
- Wataru Kagawa‡,**,
- Takashi Kinebuchi‡,
- Mitsuyoshi Yamazoe‡,
- Shigeyuki Yokoyama‡,§,**§§ and
- Takehiko Shibata‖,¶§§
- From the ‡RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan, the §Cellular Signaling Laboratory, RIKEN Harima Institute at SPring-8, 1-1-1 Kohto, Mikazuki-cho, Sayo, Hyogo 679-5143, Japan, the ‖Cellular and Molecular Biology Laboratory, RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan, ¶Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Corporation, 1637 Yana, Kisarazu, Chiba 292-0812, Japan, the **Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan, and ‡Faculty of Medicine, Kyoto University, Konoe Yoshida, Sakyo-ku, Kyoto 606-8501, Japan
Abstract
The Xrcc2 and Rad51D/Rad51L3 proteins, which belong to the Rad51 paralogs, are required for homologous recombinational repair (HRR) in vertebrates. TheXrcc2 and Rad51D/Rad51L3 genes, whose products interact with each other, have essential roles in ensuring normal embryonic development. In the present study, we coexpressed the human Xrcc2 and Rad51D/Rad51L3 proteins (Xrcc2 and Rad51D, respectively) inEscher- ichia coli, and purified the Xrcc2·Rad51D complex to homogeneity. The Xrcc2·Rad51D complex catalyzed homologous pairing between single-stranded and double-stranded DNA, similar to the function of the Xrcc3· Rad51C complex, which is another complex of the Rad51 paralogs. An electron microscopic analysis showed that Xrcc2·Rad51D formed a multimeric ring structure in the absence of DNA. In the presence of ssDNA, Xrcc2·Rad51D formed a filamentous structure, which is commonly observed among the human homologous pairing proteins, Rad51, Rad52, and Xrcc3·Rad51C.
- DSB
- double strand break
- HRR
- homologous recombinational repair
- ssDNA
- single-stranded DNA
- dsDNA
- double-stranded DNA
- Ni-NTA
- nickel-nitrilotriacetic acid
- BSA
- bovine serum albumin
- Received June 20, 2001.
- Revision received January 11, 2002.
- The American Society for Biochemistry and Molecular Biology, Inc.
