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Protein import into mitochondria and its disorders

Research Project

Project/Area Number 04454174
Research Category

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

Allocation TypeSingle-year Grants
Research Field Pathological medical chemistry
Research InstitutionKumamoto University

Principal Investigator

MORI Masataka  Kumamoto University, School of Medicine Professor, 医学部, 教授 (40009650)

Co-Investigator(Kenkyū-buntansha) TERADA Kazutoyo  School of Medicine Assistant Professor, 医学部, 助手 (00253724)
Project Period (FY) 1992 – 1993
Project Status Completed (Fiscal Year 1993)
Budget Amount *help
¥7,100,000 (Direct Cost: ¥7,100,000)
Fiscal Year 1993: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1992: ¥4,800,000 (Direct Cost: ¥4,800,000)
KeywordsMitochondrial biogenesis / Ornithine transcarbamylase / Precursor protein / Presequence / PBF / Cytosolic factor / ミトコンドリア蛋白質前駆体 / プレシークエンス
Research Abstract

Ornithine transcarbamylase(OTC), a mitochondrial matrix enzyme of the urea cycle, is initially synthesized on cytosolic free ribosomes as a larger precursor pOTC with an NH_2 -terminal presequence of 32 amino acid residues, pOTC is released into a cytosolic pool and is imported into the mitochondrial matrix with a half life of 1-2 min. We established an in vitro import system in which the purified recombinant pOTC was imported into isolated mitochondria. Using this system, we showed that a cytosolic protein factor(s) in the reticulocyte lysate is required for the import of the purified pOTC.
A protein factor that binds to pOTC but not to mature OTC and was named presequence binding factor(PBF), was purified from the lysate. The purified PBF migrated as a single polypeptide of about50,000 Da on SDS-PAGE.On sucrose gradients, pOTC and PBF cosedimented as a complex of 7S.The purified PBF markedly stimulated the import of the purified pOTC into the mitochondria. PBF-stimulated pOTC import w … More as further enhanced by hsp70 purified from yeast ; the hsp70 alone had little effect. Thus, PBF binds to the presequence portion of the precursor and may hold it in a transport-competent form in cooperation with hsp70. The precursor for aspartate aminotransferase and malate dehydrogenase synthesized in the PBF-depleted lysate failed to be imported into the mitochondria. Readdition of the purified PBF to the depleted lysate fully restored the import. Therefore, PBF appears to be involved in transport of a set of mitochondrial proteins having presequences. On the other hand, depletion of PBF from the lysate had little effect on the import of 3-oxoacyl-CoA thiolase that has no cleavable presequence. These results indicate that PBF-dependent and -independent pathways of mitochondrial protein import do exist. Proteins of the latter pathway including 3-oxoacyl-CoA thiolase may skip a PBF-dependent step and bind directly to the "presequence receptor" on the mitochondrial surface. cDNAs for mouse and human PBF were isolated based on partial amino acid sequences of the purified PBF.Mouse PBF was highly expressed in Escherichia coli and was purified to near homogeneity. Characterization of the recombinant PBF is underway. Less

Report

(3 results)
  • 1993 Annual Research Report   Final Research Report Summary
  • 1992 Annual Research Report
  • Research Products

    (24 results)

All Other

All Publications (24 results)

  • [Publications] Murakami,K.et al.: "Presequence binding factor(PBF)-dependent and independent import of proteins into mitochondria" J.Biol.Chem.267. 13119-13122 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Kanazawa,M.et al.: "Molecular cloning and sequence analysis of the cDNA for human mitochondrial short-chain enoyl-CoA hydratase" Enzyme & Protein. 47. 9-13 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Matsuno,F.et al.: "A novel ornithine transcarbamylase present in mycoplasma-infected myeloma cells" Enzyme & Protein. 47. 57-64 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 森正敬: "細胞内局在シグナルと疾患:総論" BIOmedica. 7(2). 16-17 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 森正敬: "タンパク質のミトコンドリア移行と異常" BIOmedica. 7(2). 31-35 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 村上薫、森正敬: "ミトコンドリア蛋白質の局在化と熱ショック蛋白質" 細胞工学. 11(3). 165-171 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Takiguchi,M.et al.: "“Frontiers and New Horizons in Amino Acid Research"" Elsevier, 705(145-150) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 森正敬: "新生化学実験講座1「タンパク質VI」" 東京化学同人, 456(383-391) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 森正敬: "新生化学実験講座15「代謝異常」" 東京化学同人, 359(20-24) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 森正敬: "新生化学実験講座15「代謝異常」" 東京化学同人, 359(24-26) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 大竹明、森正敬: "新生化学実験講座15「代謝異常」" 東京化学同人, 359(47-50) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] 村上薫、森正敬: "新生化学実験講座6「生体膜と膜輸送 下」" 東京化学同人, 910(723-733) (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Murakami, K.et al.: "Presequence binding factor(PBF) -dependent and- independent import of proteins into mitochondria" J.Biol. Chem.267. 13119-13122 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Kanazawa, M.et al.: "Molecular cloning and sequence analysis of the cDNA for human mitochondrial short-chain enoyl-CoA hydratase" Enzyme & Protein. 47. 9-13 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Matsuno, F.et al.: "A novel ornithine transcarbamylase present in mycoplasma-infected myeloma cells" Enzyme & Protein. 47. 57-64 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Takiguchi, M.et al.: Frontiers and New Horizons in Amino Acid Research. Elsevier, 705(145-150) (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Kanazawa,M.et al.: "Molecular cloning and sequence analysis of the cDNA for human mitochondrial short-chain enoyl-CoA hydratase" Enzyme & Protein. 47. 9-13 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] Matsuno,F.et al.: "A novel ornithine transcarbamylase present in mycoplasma-infected myeloma cells" Enzyme & Protein. 47. 57-64 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] K.Murakami,S.Tanase,Y.Morino,M.Mori: "Presequence biding factorーdependent and independent import of proteins into mitochondria" Journal of Biological Chemistry. 267. 13119-13122 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 村上 薫,森 正敬: "ミトコンドリアタンパク質の局在化と熱ショックタンパク質" 細胞工学. 11. 165-171 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 森 正敬: "細胞内局在シグナルと疾患:総論" Blomedica. 16. 124-125 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 森 正敬,村上 薫: "タンパク質のミトコンドリア移行と異常" Blomedica. 16. 139-143 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 森 正敬(分担執筆): "新生化学実験講座1「タンパク質VI」" 日本生化学会, 456P 9P (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] 村上 薫,森 正敬(分担執筆): "新生化学実験講座6「生体膜と膜輸送下」" 日本生化学会, 473P 11P (1992)

    • Related Report
      1992 Annual Research Report

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Published: 1992-04-01   Modified: 2016-04-21  

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