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

1998 Fiscal Year Final Research Report Summary

Analyses of molecular mechanisms of mitochondrial protein transport by using unnatural amino acids

Research Project

Project/Area Number 09044070
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research Field Structural biochemistry
Research InstitutionNagoya University

Principal Investigator

ENDO Toshiya  Nagoya University, Faculty of Science, Professor, 大学院・理学研究科, 教授 (70152014)

Co-Investigator(Kenkyū-buntansha) SCHULTZ Peter G.  Univ. of California, Dept. of Chemistry, Professor, バークレイ校, 教授
NISHIKAWA Shun-ichi  Nagoya University, Faculty of Science, Research Associate, 大学院・理学研究科, 助手 (10252222)
YOSHIHISA Toru  Nagoya University, R. C. M. S., Associate Professor, 物質科学国際研究センター, 助教授 (60212312)
Project Period (FY) 1997 – 1998
Keywordstranslocation across the membranes / mitichondria / chloroplasts / suppressor tRNA method / photocrosslinking / unnatural amino acid
Research Abstract

Most mitochondrial proteins are synthesized as precursor proteins in the cytosol and imported into mitochondria with the aid of protein translocation machineries in the outer and the inner membranes called the TOM complex and the TIM complex, respectively. In the present study, artificially aminoacylated suppressor tRNAs were used to introduce photoreactive unnatural amino acids into model mitochondrial precursor proteins to map interactions between precursor proteins and translocation machineries along the import pathway.
A model precursor protein, pSu9-DHFR, was arrested at two distinct stages, stage A (accumulated at 0℃) and stage B (accumulated at 30℃), in the translocation across the outer membranes and interactions between the arrested precursor protein and TOM proteins were analyzed at high resolution not achieved previously. Although the stage-A and the stage-B intermediates were previously assigned to the forms bound to the cis site and the trans site of the TOM complex, respectively, the results of crosslinking indicate that the presequence of the intermediates at both stage A and stage B is already on the trans side of the outer membrane. The mature domain is unfolded and bound to Tom40 at stage B while remains folded at stage A. After dissociation from the TOM complex, translocation of the stage-B intermediate, but not of the stage-A intermediate, across the inner membrane was promoted by the intermembrane-space domain of Tom22. These results indicate that translocation of the presequence and unfolding of the mature domain are not necessarily coupled. We have also applied the approach of site-specific photocrosslinking to the process of carrier proteins transport to the mitochondrial inner membrane and of protein translocation across the chloroplast envelope membranes.

  • Research Products

    (37 results)

All Other

All Publications (37 results)

  • [Publications] T. Kanamori et al.: "Probing the environment along the protein import pathways in yeast mitochondria by site-specific photocrosslinking"Proc. Natl. Acad. Sci. USA. 94. 485-490 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M. Sugita et al.: "Organization of a large gene cluster encoding rbosomal proteins in the cyanobacterium Synechococcus sp, strain PCC6301 : comparison of gene clusters among cyanobacteria, …"Gene. 195. 73-79 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] S. Nishikawa and T. Endo: "The yeast JEM1p is a DnaJ-like protein of the endoplasmic reticulum membrane required for nuclear fusion"J. Biol. Chem.. 272. 12889-12982 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] J. Iwahashi et al.: "Analysis of the functional domain of the rat liver mitochondrial import receptor Tom20"J. Biol. Chem.. 272. 18467-18472 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 遠藤斗志也ら: "ミトコンドリアへのターゲティングと膜透過"細胞工学. 6. 1275-1281 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] S. Nishikawa and T. Endo: "Reinvestigation of the functions of the hydrophobic segment of Jem1p, a yeast endoplasmic reticulum membrane protein mediating nuclear fusion"Biochem. Biophys. Res. Commun.. 244. 785-789 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 野原哲也,遠藤斗志也: "in vitro転写・翻訳系を用いたタンパク質合成"細胞工学別冊. 9. 37-45 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M. Oka et al.: "Loss of hsp70-hsp40 chaperone activity causes abnormal nuclear distribution and aberrant microtubule formation in M-phase of Saccharomyces cerevisiae"J. Biol. Chem.. 273. 29727-29737 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y. Kubo et al.: "Two distinct mechanisms opeartein reactivation of heat-denatured proteins by the mitochondrial Hsp70/Mdj1p/Yge1p chaperone system"J. Mol. Biol.. 286. 447-464 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M. Yoshida et al.: "Dynamics and regulation of the stress response : meeting report onthe stress response snd molecular chaperones from Kyoto"Cell Stress & Chaperones. 4. 66-74 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T. Kanamori et al.: "Uncoupling of transfer of the presequence and unfolding of the mature domain in precursor translocation across the mitochondrial outer membrane"Proc. Natl. Acad. Sci. USA. 96. 3634-3639 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M. Esaki et al.: "Two distinct mechanisms drive protein translocation across the mitochondrial outer membrane in the late step of the cytochrome b_2 import pathway"Proc. Natl. Acad. Sci. USA. 96. 11770-11775 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T. Asai et al.: "Sec-dependent pathway and ΔpH-dependent pathway do not share a common translocation pore in thylakoidal protein transport"J. Biol. Chem.. 274. 20075-20078 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 吉久 徹,遠藤斗志也: "葉緑体への蛋白質輸送"蛋白質核酸酵素. 45. 139-146 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y. Abe et al.: "Structural basis of presequece recognition by the mitochondrial protein import receptor Tom20"Cell. 100. 551-560 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T. Endo(分担執筆): "Assembly of thylakoid membrane proteins in chloroplasts"Membrane Proteins : Structure, Function and Expression Control.(N. Hamasaki and K. Mihara eds) Kyushu University Press / S. Karger AG, Fukuoka / Basel. 191-197 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T. Endo(分担執筆): "Molecular Chaperones in Proteins : Structure, Function, and Mode of Action (A. Fink and Y. Goto, eds) Marcel Dekker, Inc. New York"Roles of molecular chaperones in mitochondrial protein import. 435-466 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 西川周一,遠藤斗志也(分担執筆): "酵母ラボマニュアル:酵母分子細胞生物学実験法(山本正幸,大矢禎一編)シュプリンgガーフェアラーク(東京)"オルガネラの機能解析:ミトコンドリア. 191-211 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 西川周一,遠藤斗志也(分担執筆): "新版 微生物学実験法(大和田紘一,黒岩常祥,杉村純多,高橋秀夫,徳田 元,渡辺 信編)高段者サイエンティフィック(東京)"酵母の細胞内構造体の調製法. 121-125 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 遠藤斗志也(分担執筆): "細胞内物質輸送のダイナミズム(米田悦啓,中野明彦編)シュプリンガーフェアラーク東京"葉緑体への輸送. 65-73 (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 西川周一,遠藤斗志也(分担執筆): "生物化学実験法シリーズ(江尻慎一郎,平 秀晴,堤 賢一,志村憲介編)学会出版センター"無細胞蛋白質生合成系:蛋白質生産システム. 257-264 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T. Kanamori et al.: "Probing the environment along the protein import pathways in yeast mitichondria by site-specific photocrosslinking"Proc. Natl. Acad. Sci. U. S. A.. 94. 485-490 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Sugita rt al.: "Organization of a large gene cluster encoding ribosomal proteins in the cyanobacterium Synechococcus sp. strain PCC6301"Gene. 195. 73-79 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Endo et al.: "Mitochondrial protein targeting and translocation across the membranes"Cell Tchnology. 16 (In Japanese). 1275-1281 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] J. Iwahashi et al.: "Analysis of the functional domain of the rat liver mitochondrial import receptor Tom20"J. Biol. Chem.. 272. 18467-18472 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] S. Nishikawa and T. Endo: "The yeast JEMlp is a DnaJ-like protein of the endoplasmic reticulum membrane required for nuclear fusion"J. Biol. Chem. 272. 12889-12892 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] S. Nishikawa and T. Endo: "Reinvestigation of the functions of the hydrophobic segment of Jemlp, a yeast endoplasmic reticulum membrane protein mediating nuclear fusion"Biochem. Biophys. Res. Commun.. 244. 785-789 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Oka et al.: "Loss of hsp70-hsp70 chaperone activity causes abnormal nuclear distribution and abberrant microtubule formation in M-phase of Saccharomyces cerevisiae"J. Biol. Chem.. 273. 29727-29737 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Y. Kubo et al.: "Two distinct mechanisms operate in reactivation of heat-denatured proteins by the mitochondrial Hsp70/Mdjlp/Ygelp chaperone system"J. Mol. Biol.. 286. 447-464 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Kanamori et al.: "Uncoupling of transfer of the presequence and unfolding of the mature domain in precursor translocation across the mitochondrial membrane"Proc. Natl. Acad Sci. USA. 96. 3634-3639 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Asai et al.: "Sec-dependent pathway and ΔpH-dependent pathway do not share common translocation pore in thylakoidal protein transport"J. Biol. Chem. 274. 20075-20078 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Yoshida et al.: "Dynamics and rgulation of the stress response : meeting report on the stress response and mlecular chaperones from Kyoto"Cell Stress & Chaperones. 4. 66-74 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Esaki et al.: "Two distinct mechanisms drive protein translocation across the mitochondrial outer membrane in the late step of the cytochrome bィイD22ィエD2 import pathway"Proc. Natl. Acad. Sci. USA. 96. 11770-11775 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Yoshihisaand T. Endo: "Choroplast protein import"PNE. 45 (in Japanese). 139-146 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Y. Abe et al.: "Structural basis of presequence recognition by the mitochondrial import receptor Tom20"Cell. 100. 551-560 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. endo: "Roles of molecular chaperones in mitochondrial protein import"Molecular Chaperones in Proteins : Structure, Function, and Mode of Action, A. Fink and Y. Goto, eds. (New York : Marcel Dekker, Inc.). 435-466 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Endo: "Assembly of thylakoid membrane proteins in chloroplasts"Membrane Proteins : Structure, Function and Expression Control, N. Hamasaki and K. Mihara, eds. (Basel : Karger). 191-197 (1997)

    • Description
      「研究成果報告書概要(欧文)」より

URL: 

Published: 2001-10-23  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi