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CHARGERIN---ITS TERTIARY STRUCTURE AND THE MECHANISM OF ENERGY TRANSDUCTION

Research Project

Project/Area Number 63480502
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field 代謝生物化学
Research InstitutionTOKUSHIMA UNIVERSITY

Principal Investigator

HIGUTI Tomihiko  TOKUSHIMA UNIVERSITY, PHARMACEUTICAL SCIENCES, ASSOCIATE PROFESSOR, 薬学部, 助教授 (50035557)

Project Period (FY) 1988 – 1989
Project Status Completed (Fiscal Year 1989)
Budget Amount *help
¥6,900,000 (Direct Cost: ¥6,900,000)
Fiscal Year 1989: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1988: ¥5,500,000 (Direct Cost: ¥5,500,000)
KeywordsMitochondria / Mitochondrial DNA / Energy transduction / Chargerin / ATP synthase / Proton pump / subunit b / Molecular cloning / バイオジェネシス / エネルギー変換 / チャージリン
Research Abstract

Recently we purified a hydrophobic protein named chargerin 11 from rat liver mitochondria [Higuti, T., et al. J. Biol. Chem. 263, 6772-6776 (1988)] , which was encoded by the unidentified reading frame URFA6L of mitochondrial DNA. Although chargerin IT is hydrophobic and soluble in a mixture of chloroform and methanol (2:1), it has unbalanced positive charges in its sequence. Chargerin 11 is one of the subunits of Fo of H^+-ATP synthase. Furthermore. an antibody against chargerin II inhibited energy-transduction by H^+-ATP synthase in mitoplasts in an energy-dependent fashion. suggesting that energization of H^+-ATP synthase causes a conformational change in chargerin II [Uchida. J. et al. Biochem. Biophys. Res. Commun. 165, 449-456 (1987)].
These unique features of chargerin II suggest that it has an essential role in the energy transduction by mitochondrial ATP synthase, in good accord with the charge-transfer coupling hypothesis [Higuti, T. , Mol. Cell. Biochem. 166, 37-61 (1984)].
In … More the present research project. we have obtained the following important findings.
1. The orientation of chargerin II in Fo of the ATP synthase of rat liver mitochondria was examined using antibodies against peptides of chargerin II. Results showed that its N-terminal region (about 8 amino acid residues) was exposed on the surface of the C-side of Fo, but its C-terminal and charge-cluster regions were buried in Fo.
2. By using the anisotropic inhibitors of energy transduction. which were found by us previously, we found that "an internal electric field" in H^+-transport redox complexes and ATP synthase is formed in their energized state. in good accord with the charge-transfer coupling hypothesis.
3. The contents of chargerin If in the H^+-ATP synthase purified from rat liver mitochondria and in submitochondrial particles were determined by radioimmunoassay. Results showed that the H^+-ATP synthase contained chargerin II in a molar ratio of one to one. This is the first report on the stoichiometry of the A6L-product in mitochondrial H^+-ATP synthase.
4. The subunits of H^+-ATP synthase from rat liver mitochondria were purified on a reverse-phase column. The sequences of these subunits were determined by the protein sequences. The subunit b and factor 6 were firstly found in rat. Then we synthesized the probe DNA for the subunit b and succeeded to clarify the sequence of cDNA for the import precursor of subunit b. The cDNA of subunit b contained 1,124 base pairs. The sequence contained the coding region of 42 amino acids of the import signal reptide and 214 amino acids of the mature protein. Less

Report

(3 results)
  • 1989 Annual Research Report   Final Research Report Summary
  • 1988 Annual Research Report
  • Research Products

    (30 results)

All Other

All Publications (30 results)

  • [Publications] Higuti,T.,: "A Hydrophobic Protein,Chargerin II,Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." J.Biol.Chem.263. 6772-6776 (1988)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Oda,T.,: "Orientation of Chargerin II(A6L)in the ATP synthase of Rat Liver Mitochondria Determined with Antibodies against Peptides of the Protein." Biochem.Biophys.Res.Commun.165. 449-456 (1989)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti,T.,: "A Hydrophobic Prtein,Chargerin II,Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." In:Molecular Structure,Function and,and Assembly of ATP Synthases(S.Marzuki,ed.)Plenum Press,. 271-277 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Muraguchi,M.,: "Stoichiometry of Chargerin II(A6L)in the H^+-ATP Synthase of Rat Liver Mitochondria." Biochem.Biophys.Res.Commun.,(in press). (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Yoshihara,Y.,: "H^+-ATP Synthase from Rat Liver Mitochondria.A Simple,Rapid Purification Method of The Functional Complex and Its Characterization." J.Biol.Chem.(submitted.).

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Turumi,T.,: "cDNA Sequence of the Import Precursor of the Subunit b of H^+-ATP Synthase from Rat Mitochondria." Biochemistry(submitted).

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] 樋口富彦: "生体膜における電子移動-ミトコンドリアを例として-「バイオ素子開発と設計」赤池敏宏,品川嘉也編集" ミマツデ-タシステム, 34 (1989)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] 樋口富彦: "ドットブロットとウエスタンブロット法,「細胞・抗体・遺伝子の基礎実験法」-蛋白質を中心として,堀尾武一監修(印刷中)" 南江堂, (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti, T., Negama, T., Takigawa, M., Uchida, J., Yamane, T., Asai, T., Tani, I., Oeda, K., Shimizu, M., Nakamura, K., and Ohkawa, H.: "A Hydrophobic Protein, Chargerin II, Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." J. Biol. Chem., 263, 6772-6776 (1988).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Oda, T., Futaki, S., Kitagawa, K., Yoshihara, Y., Tani, I., & Higuti, T.: "Orientation of Chargerin II (A6L) in the ATP synthase of Rat Liver Mitochondria Determined with Antibodies against Peptides of the Protein." Biochem. Biophys. Res. Commun. 165, 449-456 (1989).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti, T., Negama, T., Takigawa, M., Uchida, J., Yamane, T., Asai, T., Tani, I., Oeda, K., Shimizu, M., Nakamura, K., and Ohkawa, H.: "A Hydrophobic Protein, Chargerin II, Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." In: Molecular Structure, Function and, and Assembly of ATP Synthases (S. Marzuki, ed.) Plenum Press, pp. 271-277 (1990).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Muraguchi, M., Yoshihara, Y., Tunemitu, T., Tani, I., and Higuti, T.: "Stoichiometry of Chargerin II (A6L) in the H^+-ATP Synthase of Rat Liver Mitochondria." Biochem. Biophys. Res. Commun. (1990).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Yoshihara, Y., Nagase, H., Yamane, T., Oka, H., Tani, I., & Higuti, T.: "H^+-ATP Synthase from Rat Liver Mitochondria. A Simple, Rapid Purification Method of The Functional Complex and Its Characterization." J. Biol. Chem.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Tsurumi, C., Yoshihara, Y., Osaka, F., Yamada, F., Tani, I., Higuti, T., Shimizu, M., Oeda, K., Ohkawa, H., Toda, H., Kakuno, T., Sakiyama, F., Kumatori, M., Tanaka, K., and Ichihara, A.: "cDNA Cloning and Sequencing For The Import Precursor of Subunit B In H^+-ATP Synthase From Rat Mitochondria" Biochem. Biophys. Res. Commun. (1990).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti, T., Hirano, K., Nagase, H., and Yoshihara, Y.: "Additiveness of Tetraphenylphosphonium-dependent H^+-Transport in Mitochondria Energized by Respiratory Substrate and ATP." Manuscript.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Nagase, H., Ohsaka, F., Yoshihara, Y., Tani, I., and Higuti, T.: "Additiveness of Anilinonaphthalene Sulphonate-Response to the Energization of Submitochondrial Particles Caused by Respiratory Substrate and ATP." Manuscript.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti, T., Harada, K., Tunemitu, T., Higuchi, S., Maeda, Y., and Matumoto, K.: "A Highly Active Protein Synthesis in a Novel Type of Mitochondria found from Rat Liver." Manuscript.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Higuti,T.: "A Hydrophobic Protein,Chargerin II,Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." J.Biol.Chem.263. 6772-6776 (1988)

    • Related Report
      1989 Annual Research Report
  • [Publications] Oda,T.: "Orientation of Chargerin II(A6L)in the ATP synthase of Rat Liver Mitochondria Detemined with Antibodies against Peptides of the Protein." Biochem.Biophys.Res.Commun.165. 449-456 (1989)

    • Related Report
      1989 Annual Research Report
  • [Publications] Higuti,T.: "A Hydrophobic Protein,Chargerin II,Purified from Rat Liver Mitochondria Is Encoded in the Unidentified Reading Frame A6L of Mitochondrial DNA." In:Molecular Structure,Function and,and Assembly of ATP Synthases(S.Marzuki,ed.)Plenum Press. 271-277 (1990)

    • Related Report
      1989 Annual Research Report
  • [Publications] Muraguchi,M.: "Stoichiometry of Chargerin II(A6L)in the H^+-ATP Synthase of Rat Liver Mitochondria." Biochem.Biophys.Res.Commun.(1990)

    • Related Report
      1989 Annual Research Report
  • [Publications] Yoshihara,Y.: "H^+-ATP Synthase from Rat Liver Mitochondria.A Simple,Rapid Purification Method of The Functional Complex and Its Characterization." J.Biol.Chem.

    • Related Report
      1989 Annual Research Report
  • [Publications] Turumi,T.: "cDNA Sequence of the Import Precursor of the Subunit b of H^+-ATP Synthase from Rat Mitochondria." Biochemistry.

    • Related Report
      1989 Annual Research Report
  • [Publications] 樋口富彦: "生体膜における電子移動ーミトコンドリアを例としてー「バイオ素子開発と設計」赤池敏宏・品川嘉也編集" ミマツデ-タシステム, 34 (1989)

    • Related Report
      1989 Annual Research Report
  • [Publications] 樋口富彦: "ドットブロットとウエスタンブロット法「細胞・抗体・遺伝子の基礎実験法」ー蛋白質を中心として堀尾武一監修" 南江堂, (1990)

    • Related Report
      1989 Annual Research Report
  • [Publications] T.Higuti et al.: J.Biol.Chem.263. 6776-6776 (1988)

    • Related Report
      1988 Annual Research Report
  • [Publications] T.Higuti et al.: Plenum Press.

    • Related Report
      1988 Annual Research Report
  • [Publications] T.Higuti et al.: J.Biol.Chem.(1989)

    • Related Report
      1988 Annual Research Report
  • [Publications] T.Higuti et al.: J.Biol.Chem.(1989)

    • Related Report
      1988 Annual Research Report
  • [Publications] 樋口富彦: "高機能システム素子としてのバイオ素子開発" ミマツデータシステム, (1989)

    • Related Report
      1988 Annual Research Report

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Published: 1988-04-01   Modified: 2016-04-21  

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