• Search Research Projects
  • Search Researchers
  • How to Use
  1. Back to previous page

Molecular biological and crystallographycal anaylsis of Clostridium perfringens iota-toxin

Research Project

Project/Area Number 14570251
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Bacteriology (including Mycology)
Research InstitutionTokushima Bunri University

Principal Investigator

SAKURAI J.  Tokusbima Bunri Univ., Fac. Pharm. Sci., Full Professor, 薬学部, 教授 (80029800)

Co-Investigator(Kenkyū-buntansha) KOBAYASHI K.  Tokusbima Bunri Univ., Fac. Pharm. Sci., Res. Associate, 薬学部, 助手 (90170315)
NAGAHAMA M.  Tokusbima Bunri Univ., Fac. Pharm. Sci., Assis.1 Professor, 薬学部, 助教授 (40164462)
Project Period (FY) 2002 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2003: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2002: ¥1,800,000 (Direct Cost: ¥1,800,000)
KeywordsClostrdium perfringens / Iota-toxin / binary toxin / ADP ribosylation / oligomer / lipid raft / endocytosis / endosome / ADPリボシル化酵素 / 結晶解析 / アミノ酸残基 / 部位特異的変異法 / ドメイン
Research Abstract

Clostridium perfringens iota-toxin is a binary toxin composed of an enzymatic component (Ia) and binding component (Ib). Ib oligomerizes to form ion-permeable channels in membranes and the oligomer induces endocyosis. To elucidate the mode of action of iota-toxin, we examined the binding and internalization of Ib using Cy3-labeled Jib. The labeled Ib was retained at the plasma membrane of MDCK cells till 60 min of incubation at 37℃, and was detected in cytoplasmic vesicles within 120 min. Treatment of the cells with methyl-β-cyclodextrin (MβCD) resulted in a marked reduction in binding to and internalization into the cells of Ib. To determine whether Ib associates with lipid rafts, MDCK cells were incubated with Ib at 4 or 37 ℃ and the Triton-insoluble membranes were fractionated by sucrose density gradient centrifugation. An Ib complex of 500 kDa was localized at 37 ℃ to the insoluble fractions that fulfilled the criteria of lipid rafts, but did not form at 4 ℃. The amount of complex in the lipid raft fraction reached a maximum after 60 min of incubation at 37 ℃ and the complex disappeared within 120 min. When the cells preincubated with Ib at 4 ℃ for 30 min were incubated at 37 ℃ for 60 min, the complex was detected in the raft fraction. Treatment of MDCK cells with Mith MβCD reduced the localization of the Tb complex to the lipid rafts and rounding of the cells induced by Ia plus Ib. When ^<125> I-labeled Ia was incubated with the cells in the presence of Ib at 37 ℃, it was localized in the lipid raft fraction. Surface plasmon resonance analysis revealed that Ia binds to the oligomer of Ib, but not the monomer. We conclude that Ib binds to a receptor in plasma membranes, then moves to lipid rafts, and Ia bound to the oligomer of Ib formed in the rafts is internalized in the cells.

Report

(3 results)
  • 2003 Annual Research Report   Final Research Report Summary
  • 2002 Annual Research Report
  • Research Products

    (27 results)

All Other

All Publications (27 results)

  • [Publications] M.Nagahama, S.Hayashi, S.Morimitsu, J.Sakurai: "Biological activities and pore-formation of Clostridium perfringens beta-toxin in HL-60 cells"J.Biol.Chem.. 278. 36934-36941 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] S.Ochi, M.Oda, M.Nagahama, J.Sakurai: "Clostridium perfringens alpha-toxin induced hemolysis of horse erythrocytes is dependent on Ca^<2+> uptake"Biochim.Biophys.Acta. 1613. 79-86 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] J.Sakurai, M.Nagahama: "Mechanism of action of Clostridium perfringens beta-toxin"Recent Res. Devel. Infection & Immunity. 1. 433-449 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] J.Sakurai, M.Nagahama, J.Hisatsune, H.Tsuge, N.Katunuma et al.: "Clostridium perfringens iota-toxin, ADP-ribosyltransferase : the structure and the mechanism of action"Advan.Enzyme.Regul.. 43. 361-377 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] H.Tsuge, M.Nagahama, H.Nishimura, N.Katunuma, J.Sakurai et al.: "Crystal structure and site-directed mutagenesis of enzymatic components from Clostridium perfringens iota-toxin"J.Mol.Biol.. 325. 471-483 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, S.Morimatsu, A.Kihara, J.Sakurai et al.: "Involvement of tachykinin receptors in Clostridium perfringens beta-toxin-induced plasma extravasation"Br.J.Pharmacol.. 138. 23-30 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] 櫻井純, 本田武司, 小熊恵二(編): "細菌毒素ハンドブック"(株)サイエンスフォーラム. 587 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, S.Hayashi, S.Morinntsu, J.Sakural.: "Biological activities and pore-formation of Clostridium perfringens beta-toxin in HL 60 cells."J. Biol. Chem.. 278. 36934-36941 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] S.Ochi, M.Oda, M.Nagahama, J.Sakurai.: "Clostridium perfringens alpha-toxin-induced hemolysis of horse erythrocytes is dependent on Ca^<2+> uptake."Biochim. Biophys. Acta,. 1613. 79-86 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] J.Sakurai, M.Nagahama.: "Mechanism of action of Clostridium perfringens beta-toxin."Recent Res. Devel. In fection & Immunity 1. 1. 433-449 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] J.Sakural, M.Nagahama, J.Hisatsune, H.Tsuge, N.Katunuma.: "Clostridium perfringens iota-toxin, ADP-ribosyltransferase: the structure and the mechanism of action."Advan. Enzyme. Regul.. 43. 361-377 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] H.Tsuge, M.Nagahama, H.Nishimura, J.Hisatsune, Y.Sakaguchi, Y.Itogawa, N.Katunuma, J.Sakurai.: "Crystal structure and site-directed mutagenesis of enzymatic component from Clostridium perfringens iota-toxin."J. Mol. Biol.. 325. 471-483 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, S.Morimitsu, A.Kihara, M.Akita, K.Setsu, J.Sakural.: "Involvement of tachykinin receptors in Clostridium perfringens beta-toxin-induced plasma extravasation."Br. J. Pharmacol.. 138. 23-30 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, M.u Mukai, S.Morimitsu, J.Sakurai.: "Role of the C-domain in the biological activities of Clostridium perfringens alpha-toxin"Microbiol. Immunol.. 46. 647-655 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, K.Nagayasu, K.Kobayashi, J.Sakurai.: "Binding component of Clostridium perfringens iota-toxin induces endocytosis in Vero cells."Infect. lmmun.. 70. 1909-1914 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] M.Nagahama, S.Hayashi, S.Morimitsu, J.Sakurai: "Biological activities and pore-formation of Clostridium perfringens beta-toxin in HL 60 cells"J.Biol.Chem.. 278. 36934-36941 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] S.Ochi, M.Oda, M.Nagahama, J.Sakurai: "Clostridium perfringens alpha-toxin-induced hemolysis of horse erythrocytes is dependent on Ca^<2+> uptake"Biochim.Biophys.Acta. 1613. 79-86 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] J.Sakurai, M.Nagahama: "Mechanism of action of Clostridium perfringens beta-toxin"Recent Res.Devel.Infection & Immunity. 1. 433-449 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] J.Sakurai, M.Nagahama, J.Hisatsune, H.Tsuge, J.Sakurai et al.: "Clostridium perfringens iota-toxin, ADP-ribosyltransferase : the structure and the mechanism of action"Advan.Enzyme.Regul.. 43. 361-377 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] H.Tsuge, M.Nagahama, H.Nishimura, N.Katunuma, J.Sakurai et al.: "Crystal structure and site-directed mutagenesis of enzymatic component from Clostridium perfringens iota-toxin"J.Mol.Biol.. 325. 471-483 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] M.Nagahama, S.Morimitsu, A.Kihara, J.Sakurai et al.: "Involvement of tachykinin receptors in Clostridium perfringens beta-toxin-induced plasma extravasation"Br.J.Pharmacol.. 138. 23-30 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] 櫻井純, 本田武司, 小熊恵二: "細菌毒素ハンドブック"(株)サイエンスフォーラム. 587 (2002)

    • Related Report
      2003 Annual Research Report
  • [Publications] M.Nagahama, M.Mukai, S.Morimitsu J.Sakurai: "Role of the C-domain in the biological activities of Clostridium perfringens alpha-toxin"Microbiology and Immunology. 46. 647-655 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] M.Nagahama, S.Morimitsu, A.Kihara, M.Akita, K.Setsu, J.Sakurai: "Involvement of tachykinin receptors in Clostridium perfringens beta-toxin-induced plasma extravasation"British Journal of Pharmacology. 138. 23-30 (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] H.Tsuge, M.Nagahama, H.Nishimura, J.Hisatsune, J.Sakurai et al.: "Crystal structure and site-directed mutagensis of enzymatic components from Clostridium perfringens iota-toxin"Journal of Molecular Biology. 325. 471-483 (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] J.Sakruai, M.Nagahama: "Mechanism of action of Clostridium perfringens beta-toxin"Recent Research Developments in Infection & Immunity. (In press).

    • Related Report
      2002 Annual Research Report
  • [Publications] J.Sakurai, M.Nagahama, J.Hisatsune, N.Katunuma, H.Tsuge: "Clostridium perfringens ι-toxin, ADP-ribosyltransferase: structure and mechanism of action"Advances in enzyme regulation. (In press).

    • Related Report
      2002 Annual Research Report

URL: 

Published: 2002-04-01   Modified: 2016-04-21  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi