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ペルオキシソーム膜ABCタンパク質の構造・機能と遺伝病

Research Project

Project/Area Number 10217203
Research Category

Grant-in-Aid for Scientific Research on Priority Areas

Allocation TypeSingle-year Grants
Review Section Biological Sciences
Research InstitutionToyama Medical and Pharmaceutical University

Principal Investigator

今中 常雄  富山医科薬科大学, 薬学部, 教授 (50119559)

Co-Investigator(Kenkyū-buntansha) 宇根 瑞穂  広島大学, 大学院・医歯薬総合研究科, 助手 (20144826)
守田 雅志  富山医科薬科大学, 薬学部, 助手 (20191033)
岡村 昭治  富山医科薬科大学, 薬学部, 助教授 (60019122)
古谷 博和  九州大学, 大学院・医学研究院, 助教授 (60253415)
Project Period (FY) 1998 – 2002
Project Status Completed (Fiscal Year 2002)
Budget Amount *help
¥47,100,000 (Direct Cost: ¥47,100,000)
Fiscal Year 2002: ¥8,900,000 (Direct Cost: ¥8,900,000)
Fiscal Year 2001: ¥9,000,000 (Direct Cost: ¥9,000,000)
Fiscal Year 2000: ¥9,400,000 (Direct Cost: ¥9,400,000)
Fiscal Year 1999: ¥9,800,000 (Direct Cost: ¥9,800,000)
Fiscal Year 1998: ¥10,000,000 (Direct Cost: ¥10,000,000)
KeywordsABCタンパク質 / トランスポーター / ペルオキシソーム / Pex19p / タンパク質発現系 / 副腎脳白質ジストロフィー / 脂肪酸代謝 / オルガネラ局在化 / 昆虫細胞発現系 / 脂質代謝 / 脂肪酸輸送 / 脂肪酸β酸化 / 極長鎖脂肪酸 / ABC蛋白 / トランスポータ / ペルオキシソーム病 / アシル-CoA輸送 / adrenoleukodystrophy
Research Abstract

ペルオキシソーム膜上のABCタンパク質の局在化機構、膜上での構造と機能、ABCタンパク質異常と疾患について解析し、以下の新たな知見を得た。
1.PMP70の翻訳時にPMP70とPex19pが結合することが、ペルオキシソーム局在化に必須であることを発見した。またこの結合には、PMP70のN末端60アミノ酸残基と膜貫通領域5、6が重要な役割を果たすことを示した。
2.ラット肝ペルオキシソームを種々の条件下8-azido[α-^<32>P]ATP、8-azido[γ-^<32>P]ATPとincubation後、抗PMP70、ALDP抗体で免疫沈降することにより、これらABCタンパク質がATP結合、加水分解することを実証した。現在、昆虫細胞発現系ならびに酵母発現系を用い、ABCタンパクの大量調製を行い、機能解析と結晶化を試みている。
3.PMP70、ALDPはラット肝ペルオキシソーム膜上で主としてホモダイマーとして存在するが、これらタンパク質はリン酸化される条件下で相互作用すること、さらにATP結合性をもつ90kDaペルオキシソーム膜タンパク質とも結合することを見出した。
4.ALDPの機能障害により、副腎脳白質ジストロフィー(ALD)が発症する。ALDPのミスセンス変異について、変異ALDPは、ALDPのペルオキシソーム膜上での機能不全、ペルオキシソーム以外のオルガネラへのmis-targeting、生合成後の細胞質での急速な分解などの運命をたどることを示した。
5.ALD繊維芽細胞の極長鎖脂肪酸β酸化の回復を指標に、フラボノイド化合物を中心に生薬成分のスクリーニングを行った。その結果、β酸化を回復させる種数種のフラボノイドを発見した。現在、繊維芽細胞の膜リン脂質構成脂肪酸組成の改善効果について検討中である。
以上、これまでの成果を、ABCタンパク質の生合成から膜上での機能発現ならびにALDとの関連性の観点からまとめる予定である。

Report

(5 results)
  • 2002 Annual Research Report
  • 2001 Annual Research Report
  • 2000 Annual Research Report
  • 1999 Annual Research Report
  • 1998 Annual Research Report
  • Research Products

    (32 results)

All Other

All Publications (32 results)

  • [Publications] Tanaka, A.R. et al.: "ATP binding/hydrolysis by and phosphorylation of peroxisomal ABC proteins PMP70 (ABCD3) and ALDP(ABCD1)"J. Biol Chem.. 227. 40142-40147 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Kashiwayama, Y. et al.: "Nucleotide-induced conformational changes of PMP70, an ATP binding cassette transporter on rat liver peroxisomal membranes"Biochem. Biophys. Res. Commun.. 291. 1245-1251 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Hashiguchi, N. et al.: "Peroxisomes are formed from complex membrane structures in PEX6-deficient CHO cells upon genetic complementation"Mol. Biol. Cell.. 13. 711-722 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] 守田雅志, 今中常雄: "ペルオキシソーム膜ABCタンパク質と副腎白質ジストロフィー"生化学. 74. 1166-1169 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Nakagawa, M.et al.: "Urinary bile alcohol profiles in healthy and cholestatic children"Clin.Chim.Acta. 314・1-2. 101-106 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Ito, R.et al.: "Temperature-sensitive phenotype of chinese hamster ovary cells defective in pex5 gene"Biochem.Biophys.Res.Commun.. 288・2. 321-327 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hashiguchi, N.et al.: "Peroxisomes are formed from complex membrane structures in PEX6-deficient CHO cells upon genetic complementation"Mol.Biol.Cell. 13・2. 711-722 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Kashiwayama, Y.et al.: "Nucleotide-induced conformational changes of PMP7O, an ATP binding cassette transporter on rat liver peroxisomal membranes"Biochem.Biophys.Res.Commun.. (in press). (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hung, Y.et al.: "Intermediate state of 3-ketoacyl-CoA thiolase in the Peroxisomal import process"Biochim.Biophys.Acta. (in press). (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Imanaka,T.: "The 70-kDa peroxisomal membrane protein (PMP70), an ATP-binding cassette transporter."Cell Biochem.Biophys.. 32. 131-138 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Fujiwara,C.: "Catalase-less peroxisomes. Implication in the milder forms of peroxisomes biogenesis disorders."J.Biol.Chem.. 275・47. 37271-37277 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Morita,M.: "Insulin-degrading enzyme exists inside of rat liver peroxisomes and degrades oxidaized proteins."Cell Struct.Funct.. 25・5. 299-305 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Imanaka, T.: "The 70-kDa peroxisomal membrane protein (PMP70), an ATP-binding cassette transporter"Cell Biochem. Biophys.. (in press). (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] 今中常雄: "ABCトランスポーター:医薬品開発のターゲット(2)"バイオサイエンスとインダストリー. 58・2. 101-104 (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Nakagawa, T.: "Peroxisomal membrane protein pmp47 is essential for the activation of middle chain fatty acid inside yeast peroxisomes and is associated with peroxisome proliferation"J. Biol. Chem.. 275・5. 3455-3461 (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Imanaka, T.: "Characterization of the 70-kDa peroxisomal membrane protein, and ATP-binding cassette transporter"J. Biol. Chem.. 274・17. 11968-11976 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Sato, R.: "Sterol regulatory element-binding protein negatively regulates microsomal triglycerid transfer protein gene transcription"J. Biol. Chem.. 274・35. 24714-24720 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Mori, M.: "Presence of phospholipid-neutral lipid complex structures in atherosclerotic lesions as detected by a novel monoclonal antibody"J. Biol. Chem.. 274・35. 24823-24837 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yamasaki, M.: "Formation of peroxisomes from peroxisomal ghosts in a peroxisome-deficient mammalian cell mutant upon complementation by protein microinjection"J. Biol. Chem.. 274・50. 35293-35296 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Itabe, H.: "Metabolism of oxidized phosphatidylcholines formed in oxidized low density lipoprotein by lecithin-cholesterol acyltransferase"J. Biochem.. 126・1. 153-161 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Okamura, S.: "Isotype-specific changes in the amount of β-tubulin RNA in synchronized tabacco BY2 cells"Cell Struct. Funct.. 24・3. 117-122 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 今中常雄(分担): "分子生物学(第3章 細胞構造)"丸善株式会社. 310 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Imanaka,T.: "Characterization of the 70-kDa peroxisomal membrane protein,an ATP-binding cassette transporter" J.Biol.Chem.(in press). (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Imanaka,T.: "The 70-kDa peroxisomal membrane protein(PMP70),an ATP-binding cassette transporter" Cell Biochem.Biophys.(in press). (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] 今中 常雄: "ABCタンパク質とペルオキシソーム病" 最新医学. 54・3. 434-440 (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Kinoshita,N.: "Newly identified chinese hamster ovary cell mutants are defective in biogenesis of peroxisomal membrane vesicles" J.Biol.Chem.273・37. 24122-24130 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Imamura,A.: "Temperature-sensitive phenotype of peroxisomal-assembly processes represent the milder forms of human peroxisome-biogenesis disorders" Am.J.Hum.Genet.62・6. 1539-1543 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Shimozawa,N.: "Genetic basis of peroxisome-assembly mutants of human,chinese hamster ovary cells and yeast" Am.J.Hum.Genet.63・6. 1898-1903 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Kawabe,Y.: "Characterization of two chinese hamster ovary cell lines expressing the COOH-terminal domains of sterol regulatory element-binding protein(SREBP)-1" Cell Struct.Funct.23・4. 187-192 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Itabe,H.: "Appearance of cross linked proteins in human atheroma and rat prefibrotic liver detected by a new monoclonal antibody" Biochim.Biophys.Acta. 1406・1. 28-39 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Takahashi,Y.: "Spatial pattern of smooth muscle differentiation is specified by the epithelium in the stomach of mouse embryo" Dev.Dyn.212・5. 448-460 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 今中 忠雄(分担): "分子生物学(第3章 細胞構造)(印刷中)" 丸善株式会社, (1999)

    • Related Report
      1998 Annual Research Report

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Published: 1998-04-01   Modified: 2018-03-28  

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