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Molecular evolution of recombinational mechanism

Research Project

Project/Area Number 07044199
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research InstitutionGraduate School of Science, Osaka University

Principal Investigator

KURAMITSU Seiki  Graduate School of Science, Osaka University, 大学院・理学研究科, 教授 (60153368)

Co-Investigator(Kenkyū-buntansha) ISAEVーIVANOV ウラジミール  ペテルスブルグ核物理学研究所, 分子遺伝学部門, 研究員
KIL Yuri V  Petersburg Nuclear Physics Institute, 分子放射線生物学部門, 研究員
LANZOV Vladislav A  Petersburg Muclear Physics Institute, 分子遺伝学部門, 部長
MASUI Ryoji  Graduate School of Science, Osaka University, 大学院・理学研究科, 助手 (40252580)
KATO Ryuichi  Graduate School of Science, Osaka University, 大学院・理学研究科, 助手 (50240833)
OGAWA Hideyuki  Graduate School of Science, Osaka University, 大学院・理学研究科, 教授 (70028207)
HIROTSU Ken  Faculty of Science, Osaka City University, 理学部, 教授 (10047269)
OSHIMA Toshihisa  Faculty of Science, Kyoto University of Education, 理学部, 教授 (10093345)
ISARV-IWANOV Vladimir V  Petersburg Nuclear Physics Institute
ISAEVーIVANOV イワノフ ウラジミール ヴ  ペテルスプルグ核物理学研究所, 分子遺伝学部門, 研究員
KIL Yuri.V.  ペテルスブルグ核物理学研究所, 分子遺伝子学部門, 研究員
ALEXSEEV And  ペテルスブルグ核物理学研究所, 分子遺伝子学部門, 研究員
伊藤 建夫  大阪大学, 理学部, 助教授 (40051817)
小川 智子  大阪大学, 理学部, 教授 (80028208)
Project Period (FY) 1995 – 1996
Project Status Completed (Fiscal Year 1996)
Budget Amount *help
¥9,200,000 (Direct Cost: ¥9,200,000)
Fiscal Year 1996: ¥4,600,000 (Direct Cost: ¥4,600,000)
Fiscal Year 1995: ¥4,600,000 (Direct Cost: ¥4,600,000)
KeywordsRecA / Rad51 / recombination / hyperthermophile / archaea / thermoresistance / evolution / meiosis / crystallization / RecA蛋白質 / 組換え機構 / クローニング / 立体構造解析 / 酵素反応機構 / 分子進化
Research Abstract

The RecA protein plays an indispensable role in genetic recombination in eubacteria. Recently, the RAD51 protein, which is homologous to the RecA protein, was found in many eukaryotes. However, functional domains and reaction mechanism of RecA/RAD51 protein remain uncertain. Since a protein from thermoresistant organisms is knows to be very stable and liable to be crystallization. Then we collected many thermoresistant microorganisms from hot springs in Russia and selected several bacterial strains, Methanosarcina sp. TS-2, Halococcus turomenicus B1734, Desulfurococcus amynoliticus, Halobacterium distributum B1733, B1739, adequate for biochemical study. Two highly conserved regions of the RecA/RAD51 proteins were chosen to synthesize oligonucleotide primers for PCR.The DNA fragments ware amplified in genomic DNAa by PCR.Using these fragments as a probe, we screened a gene bank of its genomic DNA.From the positive plaques, clones carrying the recA/RAD51 gene were selected by Southern hybridization and their nucleotide sequences ware determined. Comparison of primary structures of RecA/RAD51 proteins of archaea, eubacteria, and eukaryotes suggested that the archeal RecA/RAD51 protein may be an ancestral prototype of eubacterial and eularyotic enzymes. We also cloned the recA gene of another thermophilic bacterium, Thermus thermophilus HB8. Its gene product had fundamental activities for RecA protein. As T.thermophilus RecA protein was very stable, we succeeded in crystallization of this protein both in isolation and in the presence of DNA.Furthermore, we made truncated mutant of T.thermophilus RecA protein to prevent its aggregation at high protein concentrations. As this truncated protein was also stable, it was suitable for NMR measurements for resolving its tertiary structure.

Report

(3 results)
  • 1996 Annual Research Report   Final Research Report Summary
  • 1995 Annual Research Report
  • Research Products

    (60 results)

All Other

All Publications (60 results)

  • [Publications] T.Nakagawa and H.Ogawa: "Involvement of the MRE2 gene of yeast in formation of meiosis-specific double-strand breaks and crossover recombination through RNA splicing (in press)" Genes to Cells. 2. (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Okada,K.,Hirotsu,K.,Sato,M.,Hayashi,H.,and Kagamiyama,H.: "Three-Dimensional Structure of Escherichia coli Branched-Chain Amino Acid Aminotransferase at 2.5A Resolution (in press)" J.Biochem.121. (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mollova,E.T.,Metzler,D.E.,Kintanar,A.,Kagamiyama,H.,Hayashi,H.,hirotsu,K.and Miyahara,I.: "Use of ^<1H>-^<15>N heteronuclear multiple-quantum coherence NMR spectroscopy to study the active site of aspartate aminotranferase" Biochemistry. 36. 615-625 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Alexseyev,A.A.,Bakhlanova,I.V.,Zaitsev,E.N.and Lanzov,V.A.: "Genetic characteristics of new recA mutants of Escherichia coli K-12" J.Bacteriol.178. 2018-2024 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Namsaraev,E.A.,Lanzov,V.A.and Akhmedov,A.T.: "Partial purification and characterization of two types of homologous DNA pairing activity from rat testis nuclei" Biochim.Biophys.Acta. 1305. 172-180 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mizutani,H.,Miyahara,I.,Hirotsu,K.,Nishina,Y.,Shiga,K.,Setoyama,C.and Miura,R.: "Three-dimensional structure of porcine kidney D-amino acid oxidase at 3.0 angstrom" J.Biochem.120. 14-17 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Setoyama,S.,Miura,R.,Nishina,Y.,Shiga,K.,Mizutani,H.,Miyahara,I.and Hirotsu,K.: "Crystallization of expressed porcine kidney D-amino acid oxidase and preliminary X-ray crystallographic characterization" J.Biochem.119. 1114-1117 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Yamamoto,N.,Kato,R.,and Kuramitsu,S.: "Cloning,sequencing and expression of the uvrA gene from an extremely thermophilic bacterium,Thremus thermophilus HB8" Gene. 171. 103-106 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Kato,R.,Yamamoto,N.,Kito,K.,and Kuramitsu,S.: "ATPase activity of UvrB protein from Thermus thermophilus HB8,and its interaction with DNA" J.Biol.Chem.271. 9612-9618 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Takamatsu,S.,Kato,R.,and Kuramitsu,S.: "Mismatch DNA recognition protein from an extremely thermophilic bacterium,Thermus thermophilus HB8" Nucleic Acids Res.24. 640-648 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Okamoto,A.,Kato,R.,Masui,R.,Yamagishi,A.,Oshima,T.,and Kuramitus,S.: "An aspartate aminotransferase from an extremely thermophilic bacterium,Thermus thermophilus HB8." J.Biochem.119. 135-144 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Ogawa,H.,Johzuka,K.,Nakagawa,T.,Leem,S.H.,and Hagihara,A.H.: "Functions of the yeast meiotic recombination genes,MRE11 and MRE2" Adv.Biophys.31. 67-76 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mikawa,T.,Masui,R.,Ogawa,T.,Ogawa,H.and Kuramitsu,S.: "N-terminal 33 amino acid residues of Escherichia coli RecA protein contribute to its self-assembly" J.Mol.Biol.250. 471-483 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Johzuka.,K.and Ogawa,H.: "Interaction of Mre11 and Rad50 : two proteins required for DNA repair and meiosis-specific double-strand break formation in Saccharomyces cerevisiae" Genetics. 139. 1521-1532 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Terasawa,M.,Shinohara,A.,Hotta,Y.,Ogawa,H.and Ogawa,T.: "Localization of RecA-like recombination proteins on chromosomes of the lily at various meiotic stages" Genes Dev.9. 925-934 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Shinohara,A.,Ogawa,H.and Ogawa,T.: "Structure and function of RAD51 gene in eukaryote" Nippon Rinsho. 53. 239-249 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Kawaguchi,S.and Kuramitsu,S.: "Separation of heat-stable proteins from Thermus thermophilus HB8 by two-dimensional electrophoresis" Electophoresis. 16. 1060-1066 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Vinogradskaya,G.R.,Goryshin,I.Yu.,Berlin,Yu.A.and Lanzov,V.A.: "Optimization of PCR-based diagnostics for human cytomegalovirus" J.Virol.Methods. 53. 103-112 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H.Ogawa: "Molecular Mechanisms of Genetic Recombination (Advances in Biophysics,Vol.31)" Elsevier, 238 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Setoyama S., Miura, R., Nishina, Y., Shiga, K., Mizutani, H., Miyahara, I.and Hirotsu, K.: "Crystallization of ecpressed porcine kidney D-amino acid oxidase and preliminary X-ray Crystallographic characterization" J.Biochem.119. 1114-1117 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Yamamoto, N., Kato, R., and Kuramitsu, S.: "Cloning, sequencing and expression of the uvrA gene from an extremely thermophilic bacterium, Thremus thermophilus HB8" Gene. 171. 103-106 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Kato, R., Yamamoto, N., Kito, K., and Kuramitsu, S.: "ATPase activity of UvrB protein from Thermus thermophilus HB8, and its interaction with DNA" J.Biol.Chem.271. 9612-9618 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Takamatsu, S., Kato, R., and Kuramitsu, S.: "Mismatch DNA recognition protein from an extremely thermophilic bacterium, Thermusthermophilus HB8" Nucleic Acids Res.24. 640-648 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Okamoto, A., Kato, R., Masui, R., Yamagishi, A., Oshima, T., and Kuramitsu, S.: "An aspartate aminotransferase from an extremely thermophilic bacterium, Thermus thermophilus HB8." J.Biochem.119. 135-144 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Ogawa, H., Johzkuka, K., Nakagawa, T., Leem, S.H.and Hagihara, A.H.: "Functions of the yeast meiotic recombination genes, MRE11 and MRE2" Adv.Biophys.31. 67-76 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mikawa, T., Masui, R., Ogawa.T., Ogawa, H., and Kuramitsu, S.: "N-terminal 33 amino acid residues of Escherichia coli RecA protein contribute to ist self-assembly" J.Mol.Biol.250. 471-483 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Johzuka, K.and Ogawa, H.: "Interaction of Mre11 and Rad50 : two proteins required for DNA repair and meiosis-specific double-strand break formation in Saccharomyces cerevisiae" Genetics. 139. 1521-1532 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Terasawa, M., Shinohara, A., Hotta, Y., Ogawa, H.and Ogawa, T.: "Localization of RecA-like recombination proteins on chromosomes of the lily at various meiotic stages" Genes Dev.9. 925-934 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Shinohara, A., Ogawa, H.and Ogawa, T.: "Structure and function of RAD51 gene in eukaryote" Nippon Rinsho. 53. 239-249 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Kawaguchi, S.and Kuramitsu, S.: "Separation of heat-stable proteins from Thermus thermophilus HB8 by two-dimensional electrophoresis" Electophoresis. 16. 1066-1066 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mollova, E.T., Metzler, D.E., Kintanar.A., Kagamiyama, H., Hayashi, H., Hirotsu, K.and Miyahara, I.: "Use of ^1H-^<15>N heteronuclear multiple-quantum coherence NMR spectroscopy to study the active site of aspartate aminotranferase" Biochemistry. 36. 615-625 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Nakagawa and H.Ogawa: "Involvement of the MRE2 gene of yeast in formation of meiosis-specific doublestrand breaks and corossover recombination through RNA splicing (in press)" Genes to Cells. 2 (in press). (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Okada, K., Hirotsu, K., Sato, M., Hayashi, H., and Kagamiyama, H.: "Three-Dimensional Structure of Escherichia coli Branched-Chain Amino, Acid Aminotransferase at 2.5A Resolution (in press)" J.Biochem.121. (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Alexseyev, A.A., Bakhlanova, I.V., Zaitsev.E.N.and Lanzov, V.A.: "Genetic characteristics of new recA mutants of Escherichia coli K-12" J.Bacteriol.178. 2018-2024 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Namsaraev.E.A., Lanzov, V.A.and Akhmedov, A.T.: "Partial purification and characterization of two types of homologous DNA pairing activity from rat testis nuclei" Biochim Biophys.Acta. 1305. 172-180 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Mizutani, H., Miyahara, I., Hirotsu, K,, K.Nishina, Y., Shiga, K., Setoyama.C.and Miura, R.: "Three-dimensional structure of porcine kidney D-amino acid oxidase at 3.0 angstrom" J.Biochem.120. 14-17 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H.Ogawa: Elsevier. Molecular Mechanisms of Genetic Recombination (Advance in Biophysics.Vol.31), 1-238 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Nakagawa and H.Ogawa: "Involvement of the MRE2 gene of yeast in formation of meiosis-specific double-strand breaks and crossover recombination through RNA splicing(in press)" Genes to Cells. 2. (1997)

    • Related Report
      1996 Annual Research Report
  • [Publications] Okada,K.,Hirotsu,K.,Sato,M.,Hayashi,H.,and Kagamiyama,H.: "Three-Dimensional Structure of Escherichia coli Branched-Chain Amino Acid Aminotransferase at 2.5A Resolution(in press)" J.Biochem.121. (1997)

    • Related Report
      1996 Annual Research Report
  • [Publications] Mollova,E.T.,Metzler,D.E.,Kintanar,A.,Kagamiyama,H.,Hirotsu,K.and Miyahara,I.: "Use of ^1H-^<15>N heteronuclear multiple-quantum coherence NMR spectroscopy to study the active site of aspartate aminotranferase" Biochemistry. 36・2. 615-625 (1997)

    • Related Report
      1996 Annual Research Report
  • [Publications] Alexseyev,A.A.,Bakhlanova,I.V.,Zaitsev,E.N.and Lanzov,V.A.: "Genetic characteristics of new recA mutants of Escherichia coli K-12" J.Bacteriol.178・7. 2018-2024 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Namsaraev,E.A.,Lanzov,V.A.and Akhmedov,A.T.: "Partial purification and characterization of two types of homologous DNA pairing activity from rat testis nuclei" Biochim.Biophys.Acta. 1305・3. 172-180 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Mizutani,H.,Miyahara,I.,Hirotsu,K.Nishina,Y.,Shiga,K.,Setoyama,C.and Miura,R.: "Three-dimensional structure of porcine kidney D-amino acid oxidase at 3.0 angstrom" J.Biochem.120・1. 14-17 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Setoyama,S.,Miura,R.,Nishina,Y.,Shiga,K.,Mizutani,H.,Miyahara,I.and Hirotsu,K: "Crystallization of expressed porcine kidney D-amino acid oxidase and preliminary X-ray crystallographic characterization" J.Biochem.119・10. 1114-1117 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Yamamoto,N.,Kato,R.,and Kuramitsu,S.: "Cloning sequencing and expression of the uvrA gene from an extremely thermophilic bacterium,Thremus thermophilus HB8" Gene. 171・1. 103-106 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Kato,R.,Yamamoto,N.,Kito,K.,and Kuramitsu,S.: "ATPase activity of UvrB protein from Thermus thermophilus HB8,and its interaction with DNA" J.Biol.Chem.271・16. 9612-9618 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Takamatsu,S.,Kato,R.,and Kuramitsu,S.: "Mismatch DNA recognition protein from an extremely thermophilic bacterium,Thermus thermophilus HB8" Nucleic Acids Res.24・4. 640-648 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Okamoto,A.,Kato,R.,Masui,R.,Yamagishi,A.,Oshima,T.,and Kuramitsu,S.: "An aspartate aminotransferase from an extremely thermophilic bacterium,Thermus thermophilus HB8." J.Biochem.119・1. 135-144 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Kawaguchi,S.: "Separation of Heat-Stable Proteins from Thermus thermophilus HB8 by Two-Dimensional Electrophoresis" Electophoresis. 16. 1060-1066 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Mikawa,T.: "N-Terminal 33 Amino Acid Residues of Escherichia coli RecA Protein Contribute to Its Self-Assembly" J.Mol.Biol.250. 471-483 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Vinogradskaya,G.R.: "Optimization of PCR-based diagnostics for human cytomegalovirus" J.Virol.Methods. 53. 103-112 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Namsaraev,E.A.: "Recombination Properties of a Modified RecA Protein from Pseudomonas aeruginosa" Molecular Biology. 29. 460-469 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Alexeev,A.A.: "RecA protein in Escherichia coli : deficiency in C-terminal subdoman affects the protein multimerization" Doklady Acad.Nauk Russia. 345. 268-271 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Okamoto,A.: "An Aspartate Aminotransferase from an Extremely Thermophilic Bacterium,Thermus thermophilus HB8" J.Biochem.119. 135-144 (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Takamatsu,S.: "Mismatch DNA Recognition Protein from an Extremely Thermophilic Bacterium,Thermus thermophilus HB8" Nucleic Acids Res.24. 640-648 (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Yamamoto,N.: "Cloning,Sequencing and Expression of the uvrA Gene from an An Extremely Thermophilic Bacterium,Thremus thermophilus HB8" Gene,. (in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kato,R.: "ATPase Activity of UvrB Protein from Thermus thermophilus HB8,and Its Interaction with DNA" J.Biol.Chem.(in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Alexeev,A.A.: "RecA Protein in Escherichia coli : Deficiency in C-Terminal Subdoman Affects the Protein Multimerization" Doklady Acad.Nauk Russia. 345. 268-271 (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Alexeev,A.A.: "Genetic Characteristics of New recA Mutants of Escherichia coli K-12" J.Bacteriol.(in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kudrova,M.G.: "PCR-Based Molecular Diagnostic Procedure for Revealing Mycobacterium tuberculosis complex : Visualisation of the Result with the Help of Avidin-Biotin System" Biotechnology. (in press). (1996)

    • Related Report
      1995 Annual Research Report

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Published: 1995-04-01   Modified: 2016-04-21  

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