Role of the N-terminal Domain of the Human DMC1 Protein in Octamer Formation and DNA Binding (2005)

The DMC1 protein, a eukaryotic homologue of RecA that shares significant amino acid identity with RAD51, exhibits two oligomeric DNA binding forms, an octameric ring and a helical filament. In the crystal structure of the octameric ring form, the DMC1 N-terminal domain (1–81 amino acid residues) was highly flexible, with multiple conformations. On the other hand, the N-terminal domain of Rad51 makes specific interactions with the neighboring ATPase domain in the helical filament structure. To gain insights into the functional role of the N-terminal domain of DMC1, we prepared a deletion mutant, DMC1-(82–340), that lacks the N-terminal 81 amino acid residues from the human DMC1 protein. Analytical ultracentrifugation experiments revealed that, whereas full-length DMC1 forms a octamer, DMC1-(82–340) is a heptamer. Furthermore, DNA binding experiments showed that DMC1-(82–340) was completely defective in both single-stranded and double-stranded DNA binding activities. Therefore, the N-terminal domain of DMC1 is required for the formation of the octamer, which may support the proper DNA binding activity of the DMC1 protein. The DMC1 protein, a eukaryotic homologue of RecA that shares significant amino acid identity with RAD51, exhibits two oligomeric DNA binding forms, an octameric ring and a helical filament. In the crystal structure of the octameric ring form, the DMC1 N-terminal domain (1–81 amino acid residues) was highly flexible, with multiple conformations. On the other hand, the N-terminal domain of Rad51 makes specific interactions with the neighboring ATPase domain in the helical filament structure. To gain insights into the functional role of the N-terminal domain of DMC1, we prepared a deletion mutant, DMC1-(82–340), that lacks the N-terminal 81 amino acid residues from the human DMC1 protein. Analytical ultracentrifugation experiments revealed that, whereas full-length DMC1 forms a octamer, DMC1-(82–340) is a heptamer. Furthermore, DNA binding experiments showed that DMC1-(82–340) was completely defective in both single-stranded and double-stranded DNA binding activities. Therefore, the N-terminal domain of DMC1 is required for the formation of the octamer, which may support the proper DNA binding activity of the DMC1 protein. Recombination is the exchange or transfer of information between DNA molecules, and it occurs in all organisms. During meiosis, homologous recombination takes place as part of the normal meiotic process in order to generate genetic diversity. Through homologous recombination in meiosis, the genes from each parent recombine at frequencies 100- to 1000-fold higher than those of vegetative cells, leading to different genomic signatures (1.Kleckner N. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 8167-8174Crossref PubMed Scopus (355) Google Scholar, 2.Roeder G.S. Genes Dev. 1997; 11: 2600-2621Crossref PubMed Scopus (687) Google Scholar). In meiotic homologous recombination, double-stranded breaks are introduced by the Spo11 protein, and 3′-single-stranded tails are produced at the double-stranded break sites (3.Keeney S. Giroux C.N. Kleckner N. Cell. 1997; 88: 375-384Abstract Full Text Full Text PDF PubMed Scopus (1290) Google Scholar, 4.Romanienko P.J. Camerini-Otero R.D. Mol. Cell. 2000; 6: 975-987Abstract Full Text Full Text PDF PubMed Scopus (590) Google Scholar, 5.Baudat F. Manova K. Yuen J.P. Jasin M. Keeney S. Mol. Cell. 2000; 6: 989-998Abstract Full Text Full Text PDF PubMed Scopus (533) Google Scholar). Then, the homologous pairing proteins, such as Rad51 and Dmc1, and their activators are recruited to the single-stranded tails. The Rad51 and Dmc1 proteins are eukaryotic homologues of the bacterial RecA protein that promotes homologous pairing between single-stranded DNA (ssDNA) 1The abbreviations used are: ssDNA, single-stranded DNA; dsDNA, double-stranded DNA. 1The abbreviations used are: ssDNA, single-stranded DNA; dsDNA, double-stranded DNA. and double-stranded DNA (dsDNA). Dmc1 is a meiosis-specific factor, whereas Rad51 is required for both meiotic and mitotic homologous recombination (6.Bishop D.K. Park D. Xu L. Kleckner N. Cell. 1992; 69: 439-456Abstract Full Text PDF PubMed Scopus (971) Google Scholar, 7.Shinohara A. Ogawa H. Ogawa T. Cell. 1992; 69: 457-470Abstract Full Text PDF PubMed Scopus (1049) Google Scholar). DMC1 and RAD51 share ∼54% amino acid sequence identity (8.Masson J.Y. West S.C. Trends Biochem. Sci. 2001; 26: 131-136Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar). Like RecA, Rad51 and Dmc1 promote homologous pairing between two DNA molecules in vitro (9.Sung P. Science. 1994; 265: 1241-1243Crossref PubMed Scopus (751) Google Scholar, 10.Baumann P. Benson F.E. West S.C. Cell. 1996; 87: 757-766Abstract Full Text Full Text PDF PubMed Scopus (596) Google Scholar, 11.Gupta R.C. Bazemore L.R. Golub E.I. Radding C.M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 463-468Crossref PubMed Scopus (237) Google Scholar, 12.Li Z. Golub E.I. Gupta R. Radding C.M. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 11221-11226Crossref PubMed Scopus (128) Google Scholar, 13.Hong E.L. Shinohara A. Bishop D.K. J. Biol. Chem. 2001; 276: 41906-41912Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar, 14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, 15.Sehorn M.G. Sigurdsson S. Bussen W. Unger V.M. Sung P. Nature. 2004; 429: 433-437Crossref PubMed Scopus (151) Google Scholar, 16.Enomoto R. Kinebuchi T. Sato M. Yagi T. Shibata T. Kurumizaka H. Yokoyama S. J. Biol. Chem. 2004; 279: 35263-35272Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar). Despite these similarities, Rad51 and Dmc1 require different factors to promote homologous pairing efficiently. Factors such as Rad52, Rad54, and BRCA2 mediate the homologous pairing and strand exchange reaction promoted by Rad51 (17.Sung P. J. Biol. Chem. 1997; 272: 28194-28197Abstract Full Text Full Text PDF PubMed Scopus (449) Google Scholar, 18.Petukhova G. Stratton S. Sung P. Nature. 1998; 393: 91-94Crossref PubMed Scopus (344) Google Scholar, 19.Yang H. Jeffrey P.D. Miller J. Kinnucan E. Sun Y. Thoma N.H. Zheng N. Chen P.L. Lee W.H. Pavletich N.P. Science. 2002; 297: 1837-1848Crossref PubMed Scopus (552) Google Scholar). On the other hand, the mouse TBPIP/Hop2 protein (16.Enomoto R. Kinebuchi T. Sato M. Yagi T. Shibata T. Kurumizaka H. Yokoyama S. J. Biol. Chem. 2004; 279: 35263-35272Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar) and the yeast Hop2-Mnd1 complex (20.Chen Y.K. Leng C.H. Olivares H. Lee M.H. Chang Y.C. Kung W.M. Ti S.C. Lo Y.H. Wang A.H. Chang C.S. Bishop D.K. Hsueh Y.P. Wang T.F. Proc. Natl. Acad. Sci. U. S. A. 2004; 101: 10572-10577Crossref PubMed Scopus (87) Google Scholar) stimulate the Dmc1-dependent homologous pairing. Furthermore, a Dmc1-specific mediator, the Sae3-Mei5 complex, has been found in Saccharomyces cerevisiae and may promote the assembly of Dmc1 on the double-stranded break site (21.Tsubouchi S. Roeder G.S. Genetics. 2004; 168: 1219-1230Crossref PubMed Scopus (68) Google Scholar, 22.Hayase A. Takagi M. Miyazaki T. Oshiumi H. Shinohara M. Shinohara A. Cell. 2004; 119: 927-940Abstract Full Text Full Text PDF PubMed Scopus (95) Google Scholar), although the Schizosaccharomyces pombe Swi5-Sfr1 complex, composed of homologues of the S. cerevisiae Sae3-Mei5 complex, functions with Rad51 (23.Akamatsu Y. Dziadkowiec D. Ikeguchi M. Shinagawa H. Iwasaki H. Proc. Natl. Acad. Sci. U. S. A. 2003; 100: 15770-15779Crossref PubMed Scopus (101) Google Scholar). The S. cerevisiae and human RAD51 proteins bind DNA as helical nucleoprotein filaments (24.Ogawa T. Yu X. Shinohara A. Egelman E.H. Science. 1993; 26: 1896-1899Crossref Scopus (556) Google Scholar, 25.Benson F.E. Stasiak A. West S.C. EMBO J. 1994; 13: 5764-5771Crossref PubMed Scopus (395) Google Scholar). In contrast, the human DMC1 protein assumes an octameric ring structure, which forms a complex composed of stacked rings on DNA (26.Masson J.Y. Davies A.A. Hajibagheri N. Van Dyck E. Benson F.E. Stasiak A.Z. Stasiak A. West S.C. EMBO J. 1999; 18: 6552-6560Crossref PubMed Scopus (116) Google Scholar, 27.Passy S.I. Yu X. Li Z. Radding C.M. Masson J.Y. West S.C. Egelman E.H. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10684-10688Crossref PubMed Scopus (111) Google Scholar). Recently, electron micrographs of the human DMC1 protein showed that DMC1 forms a helical filament on ssDNA (15.Sehorn M.G. Sigurdsson S. Bussen W. Unger V.M. Sung P. Nature. 2004; 429: 433-437Crossref PubMed Scopus (151) Google Scholar). Therefore, unlike RAD51, DMC1 exhibits two oligomeric DNA binding forms: an octameric ring and a helical filament. The crystal structures of the RecA superfamily members, such as bacterial RecA, archaeal RadA, and eukaryotic Rad51 and Dmc1, revealed that whereas their monomeric topologies are highly conserved, their oligomeric states differ from protein to protein. The Escherichia coli RecA (28.Story R.M. Weber I.T. Steitz T.A. Nature. 1992; 355: 318-325Crossref PubMed Scopus (678) Google Scholar), S. cerevisiae Rad51 (29.Conway A.B. Lynch T.W. Zhang Y. Fortin G.S. Fung C.W. Symington L.S. Rice P.A. Nat. Struct. Mol. Biol. 2004; 11: 791-796Crossref PubMed Scopus (238) Google Scholar), and Methanococcus voltae RadA (30.Wu Y. He Y. Moya I.A. Qian X. Luo Y. Mol. Cell. 2004; 15: 423-435Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar) proteins all formed helical filaments with six monomers/turn in their crystal structures, but their helical pitches were different. The Pyrococcus furiosus Rad51 protein, which functions as a helical filament, formed a heptameric ring (31.Shin D.S. Pellegrini L. Daniels D.S. Yelent B. Craig L. Bates D. Yu D.S. Shivji M.K. Hitomi C. Arvai A.S. Volkmann N. Tsuruta H. Blundell T.L. Venkitaraman A.R. Tainer J.A. EMBO J. 2003; 22: 4566-4576Crossref PubMed Scopus (220) Google Scholar). The crystal structure of the DMC1 protein revealed a highly symmetrical octameric ring (14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar). These structures suggest a dynamic oligomeric property of the RecA superfamily in which these proteins interconvert between rings and helical filaments. Rad51 and Dmc1 are composed of two domains: the highly conserved ATPase domain, which makes up the core oligomeric structure, and the N-terminal domain, which is not conserved in the bacterial RecA protein. The ATPase domain is important for DNA binding and ATP hydrolysis and hence is also a core catalytic domain. The N-terminal domain of the human RAD51 protein physically contacts both ssDNA and dsDNA, as determined in previous NMR titration experiments and mutational analyses (32.Aihara H. Ito Y. Kurumizaka H. Yokoyama S. Shibata T. J. Mol. Biol. 1998; 290: 495-504Crossref Scopus (115) Google Scholar). Furthermore, the N-terminal domain of Rad51 specifically interacts with the neighboring ATPase domain in the helical filament structure (29.Conway A.B. Lynch T.W. Zhang Y. Fortin G.S. Fung C.W. Symington L.S. Rice P.A. Nat. Struct. Mol. Biol. 2004; 11: 791-796Crossref PubMed Scopus (238) Google Scholar, 30.Wu Y. He Y. Moya I.A. Qian X. Luo Y. Mol. Cell. 2004; 15: 423-435Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar). By contrast, the N-terminal domain of DMC1 did not stably associate with the ATPase domain of the neighboring subunit in the octameric ring, and its functional role is unknown. In the present study, a DMC1 mutant that lacks the N-terminal domain, DMC1-(82–340), was purified. Biochemical analyses revealed that the N-terminal domain of DMC1 is important for the DMC1 octamer formation and DNA binding. On the basis of these results, the functional role of the N-terminal domain of DMC1 is discussed. Protein Purification—The full-length DMC1 protein was purified as described previously (14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar). The DMC1-(82–340) gene was inserted into the pET-15b plasmid (Novagen), and the protein was overexpressed in the E. coli strain BL21(DE3) Codon Plus (Stratagene) as an N-terminal hexahistidine-tagged protein. DMC1-(82–340) was purified from a 10-liter Luria-Bertani culture incubated at 30 °C. When the A600 of the culture was 0.4–0.6, the protein expression was induced by adding isopropyl-1-thio-β-d-galactopyranoside to a final concentration of 1 mm. The cells were harvested after an overnight incubation and were lysed by sonication in buffer A (50 mm Tris-HCl buffer (pH 8.0) containing 0.5 m NaCl, 10 mm 2-mercaptoethanol, 10% glycerol, and protease inhibitors (Complete EDTA-free; Roche Applied Science)) on ice. The cell lysate was centrifuged at 27,700 × g for 20 min, and the supernatant was gently mixed by the batch method with 4 ml of nickel-nitrilotriacetic acid-agarose beads (Qiagen) for 1 h. The protein-bound beads were packed into an Econo-column (Bio-Rad) and were washed with 30 column volumes of buffer A containing 5 mm imidazole. The DMC1-(82–340) protein was eluted in a 20-column volume linear gradient of 5–300 mm imidazole in buffer A. The peak fractions, which predominantly contained DMC1-(82–340), were collected, and thrombin protease (5 units/mg of DMC1-(82–340); Amersham Biosciences) was added to cleave off the His tag. The thrombin-containing fractions were immediately dialyzed overnight at 4 °C in buffer B (50 mm Tris-HCl buffer (pH 8.0) containing 0.2 m KCl, 0.5 mm EDTA, 10 mm 2-mercaptoethanol, and 10% glycerol). The protein, which now lacked the His tag, was subjected to chromatography on a 4-ml Heparin-Sepharose (Amersham Biosciences) column. The column was washed with 20 column volumes of buffer B, and the protein was eluted with a 20-column volume linear gradient of 0.2-1.2 m KCl in buffer B. The DMC1-(82–340) protein was eluted in a sharp peak at ∼0.5 m KCl. The protein concentration was determined with a Bio-Rad protein assay kit with bovine serum albumin (Pierce) as the standard. Crystallization of DMC1-(82–340)—DMC1-(82–340) crystals were grown by the hanging drop method at 20 °C. The hanging drop was formed by adding 1 μl of DMC1-(82–340) (concentrated to 8 mg/ml) to 1 μl of the m after and the × × after 1 the DMC1-(82–340) crystals were harvested in a containing 5 m and were in a of The of the crystal was at the for DNA reaction contained final buffer of 20 mm Tris-HCl (pH 1 mm bovine serum 1 mm mm and The of DMC1 were incubated with 1 ssDNA for at °C. The were with for 20 The were by in buffer at for and were by of the and were a the binding 10 with was incubated with the of DMC1 for 5 at °C. The were by in buffer at for and were by DNA are in of for were by the of DMC1 with 1 ssDNA for 5 the DNA concentration of 30 was added with concentration of 10 the reaction were incubated for 10 min, 1 μl of by 1 μl of was and the were incubated for The were by in buffer at for and were by of the Analytical experiments were in a DMC1 and DMC1-(82–340) 0.5 mg/ml) were in a with a The protein was dialyzed 10 mm Tris-HCl buffer (pH 8.0) containing mm KCl, 0.5 mm EDTA, mm 2-mercaptoethanol, and 10% were after of at and 20 °C. the a specific volume of and a of were of the DMC1-(82–340) protein at a concentration were on a experiments were in a buffer containing 20 mm (pH and mm KCl. of the DMC1 N-terminal DMC1 protein of two the N-terminal and ATPase the electron in previous of the full-length DMC1 protein (14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar). in the electron of the ATPase domain of Dmc1 acid residues was In contrast, the electron of the N-terminal domain acid residues was not The of the electron was not the of protein as by the of the DMC1 crystal Therefore, these suggest that the N-terminal domain of DMC1 exhibits or multiple conformations. and of gain insights into the functional role of the N-terminal domain of DMC1, we a deletion mutant, DMC1-(82–340), which lacks the N-terminal 81 amino acid residues The DMC1-(82–340) mutant was overexpressed in E. coli as an N-terminal hexahistidine-tagged protein and was purified by chromatography on a column. The was with thrombin protease and was purified by Heparin-Sepharose chromatography of DMC1-(82–340) of in These DMC1-(82–340) crystals were and their was at the to the by the and of the crystals and by different were the that DMC1-(82–340) that DMC1-(82–340) and DMC1-(82–340) a in the of DMC1-(82–340) in we an ultracentrifugation we previously full-length DMC1 formed of in (14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar) the of DMC1-(82–340) was which to a and not an octamer The P. furiosus Rad51 protein formed a heptameric ring structure in its crystal (31.Shin D.S. Pellegrini L. Daniels D.S. Yelent B. Craig L. Bates D. Yu D.S. Shivji M.K. Hitomi C. Arvai A.S. Volkmann N. Tsuruta H. Blundell T.L. Venkitaraman A.R. Tainer J.A. EMBO J. 2003; 22: 4566-4576Crossref PubMed Scopus (220) Google Scholar), and a heptameric ring structure of human DMC1 was by electron S.I. Yu X. Li Z. Radding C.M. Masson J.Y. West S.C. Egelman E.H. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10684-10688Crossref PubMed Scopus (111) Google Scholar). Therefore, the of may a ring structure or to those the of DMC1-(82–340) to the heptameric The of the mutant were in the of 20 to °C. in the of DMC1-(82–340) and the full-length protein were up to °C and that the of the DMC1-(82–340) is to that of the full-length DMC1 DMC1-(82–340) in DNA and the N-terminal domain of DMC1 has a role in DNA the RAD51 N-terminal domain is to bind to DNA (32.Aihara H. Ito Y. Kurumizaka H. Yokoyama S. Shibata T. J. Mol. Biol. 1998; 290: 495-504Crossref Scopus (115) Google Scholar). To we a assay with A ssDNA and were used as in the DNA binding in A and B, full-length DMC1 to both ssDNA and DMC1-(82–340) was completely defective in both ssDNA and binding A and The DMC1-(82–340) mutant was also defective in homologous pairing between a ssDNA and a homologous dsDNA, of its defective DNA binding Therefore, the N-terminal domain of DMC1 an role in DNA binding as as in octamer of Dmc1 and Rad51 to the dynamic oligomeric of these Rad51 to DNA as helical filaments. The crystal structures of the S. cerevisiae Rad51 protein and the M. voltae RadA protein, an archaeal revealed that the N-terminal domain of Rad51 specifically interacts with the ATPase domain of the neighboring subunit in the helical filament (29.Conway A.B. Lynch T.W. Zhang Y. Fortin G.S. Fung C.W. Symington L.S. Rice P.A. Nat. Struct. Mol. Biol. 2004; 11: 791-796Crossref PubMed Scopus (238) Google Scholar, 30.Wu Y. He Y. Moya I.A. Qian X. Luo Y. Mol. Cell. 2004; 15: 423-435Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar). These that the N-terminal domain of Rad51 an important role in the filament In contrast, the N-terminal domain of the DMC1 octameric ring was not in the crystal structure, that it is or has multiple in the octameric ring structure (14.Kinebuchi T. Kagawa W. Enomoto R. Tanaka T. Miyagawa K. Shibata T. Kurumizaka H. Yokoyama S. Mol. Cell. 2004; 14: 363-374Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, In the present study, we found that DMC1-(82–340) formed a and not an octamer, that the N-terminal domain is for the DMC1 octameric ring its Therefore, the N-terminal domain may in the structure formation by the Rad51 and DMC1 it has been that DMC1 forms helical nucleoprotein filaments in to the stacked octameric rings on ssDNA (15.Sehorn M.G. Sigurdsson S. Bussen W. Unger V.M. Sung P. Nature. 2004; 429: 433-437Crossref PubMed Scopus (151) Google Scholar, J.Y. Davies A.A. Hajibagheri N. Van Dyck E. Benson F.E. Stasiak A.Z. Stasiak A. West S.C. EMBO J. 1999; 18: 6552-6560Crossref PubMed Scopus (116) Google Scholar, 27.Passy S.I. Yu X. Li Z. Radding C.M. Masson J.Y. West S.C. Egelman E.H. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10684-10688Crossref PubMed Scopus (111) Google Scholar). The amino acid of the human RAD51 and DMC1 N-terminal are highly Therefore, an to that between the N-terminal domain and the ATPase domain in the Rad51 filament may place between the in the DMC1 helical filaments. that the N-terminal domain of DMC1 functions in the from ring to filament and a as in In the the may by ATP binding. In the DMC1 octameric ring structure, the site is at the and the binding of ATP is to at the in the structure has been in proteins, RecA Yu X. S. H. Egelman E.H. 2003; 11: Full Text Full Text PDF PubMed Scopus Google Scholar), gene 4 protein T. Cell. 2000; 101: Full Text Full Text PDF PubMed Scopus Google Scholar), and D. R. W. D. Zhang R. J.A. E. A. G. Chen Nature. 2003; PubMed Scopus Google Scholar). may the formation of a DMC1 helical filament. the N-terminal domain of DMC1 may with the ATPase domain of DMC1 and the helical filament from ring to filament may place on both oligomeric states of DMC1 been to bind DNA. the DMC1 helical filament is an for its activity (15.Sehorn M.G. Sigurdsson S. Bussen W. Unger V.M. Sung P. Nature. 2004; 429: 433-437Crossref PubMed Scopus (151) Google Scholar), the DMC1 octameric ring may a DNA binding that is for the for homologous recombination on the double-stranded break a between the crystal structures of the human DMC1 protein and the S. cerevisiae Rad51 protein that the of DMC1 is at the in both the octameric ring and the In the crystal structure of the DMC1 octameric ring, the in the ATPase domain forms a with the in the neighboring ATPase domain. On the other hand, in the crystal structure of the S. cerevisiae Rad51 filament, to the of in the ATPase domain interacts with the in the neighboring N-terminal domain (29.Conway A.B. Lynch T.W. Zhang Y. Fortin G.S. Fung C.W. Symington L.S. Rice P.A. Nat. Struct. Mol. Biol. 2004; 11: 791-796Crossref PubMed Scopus (238) Google Scholar). Therefore, in the DMC1 helical filament, the is a that interacts with the neighboring N-terminal domain, the of the S. cerevisiae Rad51 filament. In the present study, we also found that DMC1-(82–340) was completely defective in both ssDNA and binding activities. A for these is that the N-terminal domain a role in DNA RAD51 (32.Aihara H. Ito Y. Kurumizaka H. Yokoyama S. Shibata T. J. Mol. Biol. 1998; 290: 495-504Crossref Scopus (115) Google Scholar). is that the heptameric by the of the N-terminal domain of DMC1 is in DNA binding. In the octameric ring, the amino acid residues for DNA binding were the which is to a DNA By contrast, the that the is to a DNA. Dmc1 and Rad51 are to in meiotic cells, and the of gene to in meiotic recombination (6.Bishop D.K. Park D. Xu L. Kleckner N. Cell. 1992; 69: 439-456Abstract Full Text PDF PubMed Scopus (971) Google Scholar, 7.Shinohara A. Ogawa H. Ogawa T. Cell. 1992; 69: 457-470Abstract Full Text PDF PubMed Scopus (1049) Google Scholar). Dmc1 and Rad51 are to that the Dmc1-specific functions are on the octameric ring of Dmc1, is found with The octameric ring may a role in homologous pairing. The octameric ring may also for the by such as the complex and the protein, which were to specifically with Dmc1, but not with Rad51 (15.Sehorn M.G. Sigurdsson S. Bussen W. Unger V.M. Sung P. Nature. 2004; 429: 433-437Crossref PubMed Scopus (151) Google Scholar, S. Roeder G.S. Genetics. 2004; 168: 1219-1230Crossref PubMed Scopus (68) Google Scholar, 22.Hayase A. Takagi M. Miyazaki T. Oshiumi H. Shinohara M. Shinohara A. Cell. 2004; 119: 927-940Abstract Full Text Full Text PDF PubMed Scopus (95) Google Scholar). analyses of the functional role of the octameric ring are required to the of Dmc1 in meiotic R. Enomoto and A. for