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Analysis of Activation Mechanism of Calpain on the Basis of its Tertiary Structure

Research Project

Project/Area Number 12308032
Research Category

Grant-in-Aid for Scientific Research (A)

Allocation TypeSingle-year Grants
Section一般
Research Field Structural biochemistry
Research InstitutionTokyo Metropolitan Institute of Gerontology

Principal Investigator

SUZUKI Koichi  Tokyo Metropolitan Institute of Gerontology, Director, 所長 (80011948)

Co-Investigator(Kenkyū-buntansha) SORIMACHI Hiroyuki  University of Tokyo, Graduate School of Agricultural and Life Sciences Department of Applied Biological Chemistry,, 農学生命科学研究科・応用生命化学専攻, 助教授 (10211327)
INOMATA Mitsushi  Tokyo Metropolitan Institute of Gerontology, Division of Molecular Biology, Department of Protein Biochemistry, Researcher, 分子生物学研究系・蛋白質生化学部門, 研究員 (30142649)
IWASHITA Yoshiko  Tokyo Metropolitan Institute of Gerontology, Division of Molecular Biology, Department of Protein Biochemistry, Department Head, 分子生物学研究系・蛋白質生化学部門, 室長 (50111498)
MAEDA Tatsuya  University of Tokyo, Institute of Molecular and Cellular Biosciences Associate Professor, 分子細胞生物学研究所, 助教授 (90280627)
Project Period (FY) 2000 – 2001
Project Status Completed (Fiscal Year 2001)
Budget Amount *help
¥33,900,000 (Direct Cost: ¥31,800,000、Indirect Cost: ¥2,100,000)
Fiscal Year 2001: ¥9,100,000 (Direct Cost: ¥7,000,000、Indirect Cost: ¥2,100,000)
Fiscal Year 2000: ¥24,800,000 (Direct Cost: ¥24,800,000)
KeywordsCalpain / Calcium / Protease / X ray structural analysis / Higher order structure of protein / Activation mechanism / Phopsholipids / 蛋白質の構造変化 / リン脂質
Research Abstract

Calpain is composed of two subunits, 80K and 30K, but 80K itself expresses full enzyme activity. The results of X ray analysis of calpain in the absence of Ca, show that the protease domain (II) of 80K comprising domains (I-IV) is composed of two sub-domains, II and Iib, which are quite similar to those in other cysteine proteases. However, they are separated and the proper active-site that exists between the two sub-domains is not formed. The activation mechanism corresponding to how these two sub-domains come closer to forn proper active-site by conformational changes induced by Ca, was examined. Two sub-domains are separated mainly by two kinds of interactions, interaction between I and cahnodulin domain (VI) in 30K, and interaction of lib with III. Upon binding Ca to VI and III, these interactions are disrupted resulting in large conformational changes that make two sub-domains come closer. Further, minor conformational changes induced by Ca binding to II are essential to make II active. Ill plays very important roles in this activation ; acting as the novel Ca binding sites, and also as the phopspholipids binding domain that determines cellular localization of active calpain. A novel Ca binding site was also predicted in II but its precise location is still unknown. To know the structure of Ca-bound form of calpain is essential to understand the final activation mechanism. Further experiments are now in progress along this line.

Report

(3 results)
  • 2001 Annual Research Report   Final Research Report Summary
  • 2000 Annual Research Report
  • Research Products

    (49 results)

All Other

All Publications (49 results)

  • [Publications] Masumoto, H.: "Crystallization and preliminary X-ray analysis of recombinant full-length human m-calpain"Acta Crystallogr.. D56. 73-75 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Strobl, S.: "The crystal structure of calcium-free human m-calpain suggests an electrostatic switch mechanism for activation by calcium"Proc Natl. Acad. Sci., USA. 97. 588-592 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H.: "Limb-girdle muscular dystrophy with calpain 3 (p94) gene mutations (calpainopathy)"NeuroScience News. 3. 20-27 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Kitagaki, H.: "Autolysis of calpain large subunit inducing irreversible dissociation of stoichiometric heterodimer of calpain"Biosci. Biotech. Biochem.. 64. 689-695 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Tagawa, K.: "Myopathy phenotype of transgenic mice expressing active site-mutated inactive p94 Skeletal muscle-specific calpain, the gene product responsible for limb girdle muscular dystrophy type 2A"Human Molecular Genetics. 9. 1392-1402 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Richard, I.: "Loss of calpain 3 proteolytic activity leads to muscular dystrophy and to apoptosis-associated I kappa B alpha/nuclear factor kappa B pathway perturbation in mice"J. Cell Biol.. 151. 1583-1590 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H.: "Skeletal muscle-specific calpain, p94, and connectin/titin : their physiological functions and relationship to limb-girdle muscular dystrophy type 2A"Adv. Exp. Med. Biol.. 481. 383-395 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Tompa, P.: "Domain III of calpain is a Ca^<++>-regulated lipid-binding domain"Biochem. Biophys. Res. Commun.. 280. 1333-1339 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Futai, E.: "Molecular cloning of PalBH, a mammalian homologue of the Aspergillus atypical calpain PalB"Biochim. Biophys. Acta. 1517. 316-319 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H.: "The structure of calpain"J. Biochem.. 129. 653-664 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Reverter, D.: "Structural basis for possible calcium-induced activation mechanism of calpain"Biol. Chem.. 382. 753-766 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Hata, S.: "Domain II of m-calpain is a Ca^<2+>-dependent cysteine protease"FEBS Letters. 501. 111-114 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Reverter, D.: "The structure of calcium-free m-calpain : implications for calcium activation and function"Thends Cardiovasc. Med.. 11. 222-229 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Hata, S.: "Both the conserved and the unique gene structure of stomach-specific calpains reveal processes of calpain gene evolution"J. Mol. Evol.. 53. 191-203 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Nakagawa, K.: "Dissociation of m-calpain subunits occurs after autolysis of the N-terminus of the catalytic subunit, and is not required for activation"J. Biohem.. 130. 605-611 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] 反町洋之: "肢帯型筋ジストロフィー"神経進歩. 44(2). 189-203 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] 秦 勝志: "カルパインスーパーファミリーの構造と機能"生化学. 73. 1129-1140 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] 反町洋之: "筋ジストロフィーとカルパイン"蛋白質核酸酵素. 46. 1772-1780 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Masumoto, H.: "Crystallization and preliminary X-ray analysis of recombinant full-length human m-calpain"Acta Crystallogr.. D56. 73-75 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Strobl, S.: "The crystal structure of calcium-free human m-calpain suggests an electrostatic switch mechanism for activation by calcium"Proc. Natl. Acad. Sci., USA. 97. 588-592 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H: "Limb-girdle muscular dystrophy with Calpain 3 (p94) gene mutations (calpainopathy)"Neuro Science News. 3. 20-27 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Kitagaki, H.: "Autolysis of calpain large subunit inducing irreversible dissociation of stoichiometric heterodimer of calpain"Biosci. Biotech. Biochem.. 64. 689-695 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Tagawa, K.: "Myopathy phenotype of transgenic mice expressing activ site-mutated inactive p94 skeletal muscle-specific calpain, the gene product responsible for limb girdle muscular dystrophy type 2A"Human Molecular Genetics. 9. 1392-1402 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Richard, I: "Loss of calpain 3 proteolytic activity leads to muscular dystrophy and to apoptosis-associated I kappa B alpha/nuclear factor kappa B pathway perturbation in mice"J. Cell Biol. 151. 1583-1590 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H: "Skeletal muscle-specific calpain, p94, and connectin/titin : their physiological functions and relationship to limb-girdle muscular dystrophy type 2A"Adv. Exp. Med. Biol. 481. 383-395 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Tompa, P.: "Domain III of calpain is a Ca<++>-regulated Hpid-binding domain"Biochem. Biophys. Commun.. 280. 1333-1339 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Futai, E.: "Molecular cloning of PalBH, a mammalian homologue of the Aspergillus atypical calpain PalB"Biochim. Biophys. Acta. 1517. 316-319 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Sorimachi, H.: "The structure of calpain"J. Biochem. 129. 653-664 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Reverter, D.: "Structural basis for possible calcium-induced activation mechanism of calpain"Biol. Chem.. 382. 753-766 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Hata, S: "Domain II of m-calpain is a Ca^<2+>-dependent cysteine protease"FEBS Letters. 501. 111-114 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Reverter, D.: "The structure of calcium-free nrcalpain : implications for calcium activation and function"Trends Cardiovasc. Med.. 11. 222-229 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Hata, S.: "Both the conserved and the unique gene structure of stomach-specific calpains reveal processes of calpain gene evolution"J. Mol. Evol. 53. 191-203 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2001 Final Research Report Summary
  • [Publications] Tompa, P.: "Domain III of calpain is a Ca^<++>-regulated lipid-binding domain"Biochem. Biophys. Res. Commun.. 280. 1333-1339 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Futai, E.: "Molecular cloning of PalBH, a mammalian homologue of the Aspergillus atypical calpain PalB."Biochem. Biophys. Acta. 1517. 316-319 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Sorimachi, H.: "The structure of calpain."J Biochem.. 129. 653-664 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Reverter, D.: "Structural basis for possible calcium-induced activation mechanism of calpain."Biol. Chem.. 382. 653-664 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hata, S.: "Domain II of m-calpain is a Ca^<2+>-dependent cysteine protease."FEBS Letters. 501. 111-114 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Reverter, D.: "The structure of calcium-free m-calpain implications for calcium activation and function."Trends Cardiovasc. Med.. 11. 222-229 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hata, S: "Both the conserved and the unique gene structure of stomach-specific calpains reveal processes of calpain gene evolution."J. Mol. Evol.. 53. 191-203 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Nakagawa, K.: "Dissociation of m-calpain subunits occurs after autolysis of the N-terminus of the catalytic subunit, and is not required for activation."J. Biohem.. 130. 605-611 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] 秦 勝志: "カルパインスーパーファミリーの構造と機能"生化学. 73. 1129-1140 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] 反町洋之: "筋ジストロフィーとカルパイン"蛋白質核酸酵素. 46. 1772-1780 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] 鈴木紘一: "カルパインファミリーメンバーの構造と疾患"Molecular Medicine. 39. 10-17 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Masumoto,H., et al.: "Crystallization and preliminary X-ray analysis of recombinant full-length human m-calpain."Acta Crystallogr.. D56. 73-75 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Strobl,S., et al.: "The crystal structure of calcium-free human m-calpain suggests an electrostatic switch mechanism for activation by calcium."Proc.Natl.Acad.Sci.USA. 97. 588-592 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Sorimachi,H., et al.: "Limb-girdle muscular dystrophy with calpain 3 (p94) gene mutations (calpainopathy)."NeuroScience News. 3. 20-27 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Kitagaki,H., et al.: "Autolysis of calpain large subunit inducing irreversible dissociation of stoichiometric heterodimer of calpain."Biosci.Biotech.Biochem.. 64. 689-695 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Tagawa,K., et al.: "Myopathy phenotype of transgenic mice expressing active site-mutated inactive p94 skeletal muscle-specific calpain, the gene product responsible for limb girdle muscular dystrophy type 2A."Human Molecular Genetics. 9. 1392-1402 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Xiang-Ping H., et al.: "Identification of a glutamic acide and aspartic acid residue essential for catalytic activity of asdpergillopepsin II, a non-pepsin type acid proteinase."J.Biol.Chem.. 275. 26607-26614 (2000)

    • Related Report
      2000 Annual Research Report

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Published: 2000-04-01   Modified: 2016-04-21  

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