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Elucidation of Ubiquinone-binding Site of Membrane-bound Quinoproteins in Oxidative Bacteria

Research Project

Project/Area Number 06660113
Research Category

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

Allocation TypeSingle-year Grants
Research Field 応用微生物学・応用生物化学
Research InstitutionYamaguchi University

Principal Investigator

MATSUSHITA Kazunobu  Yamaguchi University, Faculty of Agriculture, Department of Biological Chemistry, Professor, 農学部, 教授 (50107736)

Co-Investigator(Kenkyū-buntansha) TOYAMA Hirohide  Yamaguchi University, Faculty of Agriculture, Department of Biological Chemistry, 農学部, 助手 (60240884)
Project Period (FY) 1994 – 1995
Project Status Completed (Fiscal Year 1995)
Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1995: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1994: ¥1,200,000 (Direct Cost: ¥1,200,000)
KeywordsUbiquinone / Quinoprotein / Glucose Dehydrogenase / Alcohol Dehydrogenase / Pyrroloquinoline quinone
Research Abstract

This research project was performed by elucidating the ubiquinone-binding sites of membrane-bound quinoproteins of oxidative bacteria, glucose dehydrogenase (GDH) and alcohol dehydrogenase (ADH), which are functioning in the glucose oxidase respiratory chain of Escherichia coli and in the alcohol oxidase respiratory chain of acetic acid bacteria, respectively. GDH is a single peptide quinoprotein which consists of membrane-spanning N-terminal region and large soluble region containing prroloquinoline quinone (PQQ) and thus catalytic site for glucose. By screening specific inhibitors to prevent the reaction of GDH with ubiquinone, we have found several intense inhibitors of capsaicins analogs. Furthermore, we have examined their inhibitory mode for GDH and also the reaction site in the glucose oxidase respiratory chain. The inhibitor spectrum was also compared between GDH and mitochondrial NADH dehydrogenase, suggesting that GDH may have a structure for ubiquinone-binding site rather di … More fferent from NADH dehydrogenase. Now, we have been searching to isolate mutants of GDH insensitive to these inhibitors. Thereafter, we will figure our the amino acid residues and the structure involved in the ubiquinone-binding site by examining DNA sequence and kinetic analysis with mutated GDH.ADH,unlike GDH,is a quinohemoprotein which consists of three different subunits, subunit I containing PQQ and heme c, subunit II having 3 hemes c, and subunit III.By separating and then reconstituting the subunits of ADH,all three subunits have been shown to be required to have ubiquinone reductase activity, and the subunit II to have the ubiquinone-binding site. Furthermore, it has been shown that ADH has ubiquinol oxidase activity as well as ubiquinone reductase activity. By examining the effect of several ubiquinone analogs as the inhibitor and also the substrate, then both sites for ubiquinone oxidation and reduction was shown to have different structure and to be located in regions different from each other within subunit II. Less

Report

(3 results)
  • 1995 Annual Research Report   Final Research Report Summary
  • 1994 Annual Research Report
  • Research Products

    (23 results)

All Other

All Publications (23 results)

  • [Publications] Hiroshi Takemura et al.: "Prosthetic Group of Aldehyde Dehydrogenase in Acetic Acid Bactria Not Pyrrolo-quinoline Quinone" Bioscience Biotechnology and Biochemisiru. 58. 2082-2083 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Hidenori Yanai et al.: "Bioelectrocatalysis by Alcohol Dehydrogenase from Acetobacter aceti Adsorbed on Bare and Chemically Modified Electrodes" Denki Kagaku. 62. 1247-1248 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Kazunobu Matsushita et al.: "Soluble and membrane-bound quinoprotein D-glucose dehydrogenase of the Acinetobacter calcoaceticus ; The binding process of PQQ to the apoenzymes" Biosci. Biotech. Biochem.59. 1548-1555 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Kazunobu Matsushita et al.: "Generation mechanism and purification of an inactive form convertible in vivo to the active form of quinoprotein alcohol dehydrogenase in Gluconobacter suboxydans" J. Bacteriol.177. 6552-6559 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Kazunobu Matsushita et al.: "Function of multiple heme c moieties in intramolecular electron transport and ubiquinone reduction in the quinohemoprotein alcohol dehydrogenase-cytochromec complex of Gluconobocter suborydans" J. Biol. Chem.271 (in press). No.9 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Kimitoshi Sakamoto et al.: "Comparison of the structural features of ubiquinone reduction sites between glucose dehydrogenase in Escherichia coli and bovine heart mitochondrial complex I" Eur. J. Biochem.(in press). (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Kazunobu Matsushita et al.: "Respiratory Chains and Bioenergetics of Acetic Acid Bacteria in “Advances in Microbial Physiology" Vol. 36, p. 247-301" Academic Press, Harcourt Brace Jovanovich, Publishers, London, 50 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] H.Takemura, T.Tsuchida, F.Yoshinaga, K.Matsushita & O.Adachi: "Prosthetic group of aldehyde dehydrogenase in acetic acid bacteria not pyrroloquinoline quinone" Biosci.Biotech.Biochem.58. 2082-2083 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] H.Yanai, K.Miki, T.Ikeda & K.Matsushita: "Bioelectrocatalysis by alcohol dehydrogenase from Acetobacter aceti adsorbed on bare and chemically modified electrodes" Denki Kagaku. 62. 1247-1248 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Matsushita, H.Toyama & O.Adachi: "Respiratory Chains and Bioenergetics of Acetic Acid Bacteria" "Advances in Microbial Physiology". Vol.36 Academic Press, London. 247-301 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Matsushita, H.Toyama, M.Ameyama, O.Adachi, A.Dewanti & J.A.Duine: "Soluble and membrane-bound quinoprotein D-glucose dehydrogenase of the Acinetobacter calcoaceticus ; The binding process of PQQ to the apoenzymes" Biosci.Biotech.Biochem.59. 1548-1555 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Matsushita, T.Yakushi, Y.Takaki, H.Toyama, & O.Adachi: "Generation mechanism and purification of an inactive form convertible in vivo to the active form of quinoprotein alcohol dehydrogenase in Gluconobacter suboxydans" J.Bacteriol.177. 6552-6559 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Matsushita, T.Yakushi, H.Toyama, E.Shinagawa & O.Adachi: "Function of multiple heme c moieties in intramolecular electron transport and ubiquinone reduction in the quinohemoprotein alcohol dehydrogenase-cytochrome c complex of Gluconobacter suboxydans" J.Biol.Chem.271, No.9 (in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] K.Sakamoto, H.Miyoshi, K.Matsushita, M.Nakagawa, J.Ikeda, M.Ohshima, O.Adachi, T.Akagi, & H.Iwamura: "Comparison of the structural features of ubiquinone reduction sites between glucose dehydrogenase in Escherichia coli and bovine heart mitochondrial complex I" Eur.J.Biochem.(in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] Hirohide Toyama et al.: "Three distinct quinoprotein alconol dehyorogenases are expressed when Pseudomoras parida is grown on different alcohols" J.Bacteriol.177. 2442-2450 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kazunobu Matsushita et al.: "Soluble and membrane-bound quinoprotein D-glucose denydrogenase of the Acinerobacter calcoaceticus ; The binding process of PQQ to the apoenzymes" Biosci.Biotech.Biochem.59. 1548-1555 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kazunobu Matsushita et al.: "Generation mechanism and puntication of an inactive form convertible in vivo to the active form of quinoprotein alcohol dehydrogenase in Gluconobacter suboxydans" J.Bacteriol.177. 6552-6559 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kazunobu Matsushita et al.: "Function of multiple heme c moieties in into amoicaluas cicotron transport and ubiquinone reduction in the quinohemoprotein alcohol dehydrogenase-cytochrome c complex of Gluconobacter suboxydans" J.Biol.Chem.271. No.9 inpress (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Kimitoshi Sakamoto et al.: "Comparison of the structural features of ubiquinone reduction sites between glucose dehydrogenase in Escherichia coli and bovine heart mitochondrial complex I" Eur.J.Biochem.(in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] Hiroshi Takemura et al.: "Prosthetic Group of Aldehyde Dehydrogenase in Acetic Acid Bacteria Not Pyrroloquinoline Quinone" Bioscience Biotechnology and Biochemistry. 58. 2082-2083 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] Mariko Sato-Watanabe et al.: "Structure-Function Studies on the Ubiquinol Oxidation Site of the Cytochrome bo Complex from Escherichia coli Using p-Benzoquinonesand Subustituted Phenols" Journal of Biological Chemistry. 269. 28899-28907 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] Hidenori Yanai et al.: "Bioelectrocatalysis by Alcohol Dehydrogenase from Acetobacter aceti Adsorbed on Bare and Chemically Modified Electrodes" Denki Kagaku. 62. 1247-1248 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] Kazunobu Matsushita et al.: "Respiratory Chains and Bioenergetics of Acetic Acid Bacteria in “Advances in Microbial Physiology"Vol.36.p.247-301" Academic Press,Harcourt Brace Jovanovich,Publishers,London, 50 (1994)

    • Related Report
      1994 Annual Research Report

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Published: 1994-04-01   Modified: 2016-04-21  

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