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Comparative biochemical studies on structure, function and physiology of catechol dioxygenases

Research Project

Project/Area Number 02454565
Research Category

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

Allocation TypeSingle-year Grants
Research Field General medical chemistry
Research InstitutionShiga University of Medical Science

Principal Investigator

NOZAKI Mitsuhiro  Shiga University of Medical Science, Professor, 医学部, 教授 (00025545)

Co-Investigator(Kenkyū-buntansha) ISHIDA Tetsuo  Shiga University of Medical Science, Assistant Professor, 医学部, 助手 (10176191)
NAKAI Chieko  Shiga University of Medical Science, Assistant Professor, 医学部, 助手 (30111892)
HORIIKE Kihachiro  Shiga University of Medical Science, Associate Professor, 医学部, 助教授 (80089870)
Project Period (FY) 1990 – 1991
Project Status Completed (Fiscal Year 1991)
Budget Amount *help
¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1991: ¥1,000,000 (Direct Cost: ¥1,000,000)
KeywordsOxygenase / Catechol 1, 2-dioxygenase / Catechol 2, 3-dioxygenase / 3, 5-Dichlorocatechol dioxygenase / Nonheme iron containing enzyme / omicron-Nitrophenol / Gene cloning / Charge transfer complex / カテコ-ル2,3ー二原子酸素添加酵素 / 非ヘム鉄酸素 / 2,4ージクロロフェノキシ酢酸 / カテコ-ル1,2-二原子酸素添加酵素 / カテコ-ル2,3-二原子酸素添加酵素 / アミノ酸配列 / 塩基配列 / oーニトロフェノ-ル / 電荷移動複合体
Research Abstract

Catechol dioxygenases are classified into two groups : intradiol and extradiol. We have previously determined the DNA sequence of an extradiol enzyme, catechol 2, 3-dioxygenase. In order to pursue comparative studies between various catechol dioxygenases, we have tried to determine the DNA sequence of intradiol enzymes.
1. we have studied the well known intradiol enzyme, catechol 1, 2dioxygenase and found for the first time the existence of 3 isozymes, au, ap, op, of this enzyme from Pseudomonas arvilla C-1. we also succeeded to clone the gene for this enzyme from Pseudomonas putida mt-2.
2. An enzyme which cleaves the benzene ring of 3, 5-dichlorocatechol has been purified to homogeneity for the first time from a Pseudoiwnad, grown with a herbicide, 2, 4-dichlorophenoxyacetic acid (2, 4-D), as the sole carbon source. The enzyme was found to be a nonheme ferric dioxygenase and to catalyze the intradiol cleavage of 3, 5-dichlorocatechol. Thus, the enzyme was designated as 3, 5-dichlorocatechol 1, 2dioxygenase. Judging from the substrate specificity, kinetic constants, and N-terminal amino acid sequence, the enzyme was distinct from other chlorocatechol dioxygenases reported previously. We have also succeeded to clone the gene for this enzyme and the determination of the DNA sequence is now underway.
3. For the elucidation of the active site structure of catechol 2, 3dioxygenase, we used o-nitrophenol, a competitive inhibitor, and found it a useful active site probe for this enzyme. Using this probe, a novel mode interaction between substrate or the analog and the enzyme has been proposed. Trials were made to substitute the iron cofactor in the active site of this enzyme by other metals, and to modify the enzyme by sulfhydryl (-SH) blocking reagents or by suicide substrata. Basic information has been obtained from these studies on this enzyme.

Report

(3 results)
  • 1991 Annual Research Report   Final Research Report Summary
  • 1990 Annual Research Report
  • Research Products

    (26 results)

All Other

All Publications (26 results)

  • [Publications] C.Nakai,K.Horiike,S.Kuramitsu,H.Kagamiayama and M.Nozaki: "Three isozymes of catechol 1,2-dioxygenese(pyrocatechase),αα,αβ,and ββ,from Pseudomonas arvilla C-1." J.Biol.Chem.205. 660-665 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K.Horiike,T.Kobayashi,T.Ishida and M.Nozaki: "〇一Nitorophenol as an active site probe for catechol 2,3-dioxygenase." Yamada Conference XXVII International Symposium on Oxygenases and Oxygen Activation,edited by S.Yamamoto,M.Nozaki,Y.Ishimura. 15-18 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] M.Koyama,T.Ishida,K.Horiike,M.Nozaki and Shimada: "Urate oxidase and copper content in the liver of macular mutant mouse,a model animal for human congenital copper deficiency,Menkes kinky hair disease." J.Nutr.Sci.Vitaminol.(1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] H.Toji,K.Horiike,T.Ishida,T.Kobayashi, M.Nozaki and M.Okamoto: "Analytical and micro-preparative high-performance gel chromatography of proteins with a short colum of prous silica-based aqueous gels:Estimation on molecular size rapid determination of ligand binding constant,and purification of S-carboxymethylated proteins for microsequencing." J.Chromatography. (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Manzoor A.Bhat,T.Ishida,K.Horiike,C.S.Vaidyanathan and M.Nozaki: "Purification of 3,5-dichlorocatechol 1,2-dioxygenase A nonheme iron dioxygenase and a key enzyme in the biodegradation of a herbicide,2,4-dichlorophenoxyacetic acid (2,4-D)from Pseudomonas cepacia CSV90" J.Biol.Chem.

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Manzoor A.Bhat,et al.: "Isolation and characterization of a new plasmid encoding genes for the degradation of 2-4dichlorophenoxyacetate(2,4,-D)from Pseudomonas Cepacia CSV90" J.Bacteriol.

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] S,Yamamoto,M.Nozaki,and Y.Ishimura(editors): "International Symposium on Oxygenases and Oxygen Activation" Yamada Science Foundation, 217 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] C. Nakai, K. Horiike, S. Kuramitsu, H. Kagamiyama and M. Nozaki: "Three isozymes of catechol 1, 2-dioxygenase (pyrocatechase), alphaalpha, alphabeta, and betabeta, from Pseudomonas arvilla C-1." J. Biol. Chem. 205. 660-665 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K. Horiike, T. Kobayashi, T. Ishida and M. Nozaki: "omicron-Nitrophenol as an active site probe for catechol 2, 3-dioxygenase." Yamada Conference XXVII International Symposium on Oxygenases and Oxygen Activation. edited by S. Yamamoto, M. Nozaki, Y. Ishimura. 15-18 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] M. Koyama, T. Ishida, K. Horiike, M. Nozaki and M. Shimada: "Urate oxidase and copper content in the liver of macular mutant mouse, a model animal for human congenital copper deficiency, Menkes'kinky hair disease" J. Nutr. Sci. Vitaminol.(1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] H. Tojo, K. Horiike, T. Ishida, T. Kobayashi, M. Nozaki and M. Okamoto: "Analytical and micro-preparative high-performance gel chromatography of proteins with a short column of porous silica-based aqueous gels : Estimation of molecular size, rapid determination of ligand binding constant, and purification of S-carboxymethylated proteins for microsequencing" J. Chromatography. (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Manzoor A. Bhat, T. Ishida, K. Horiike, C. S. Vaidyanathan and M. Nozaki: "Purification of 3, 5-dichlorocatechol 1, 2-dioxygenase, A nonheme iron dioxygenase and a key enzyme in the biodegradation of a herbicide, 2, 4-dichlorophenoxyacetic acid (2, 4-D) form Pseudomonas cepacia CSV90" J. Biol. Chem.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Manzoor A. Bhat, et al.: "Isolation and characterization of a new plasmid encoding genes for the degradation of 2, 4-dichlorophenoxyacetate (2, 4-D) from Pseudomonas cepacia CSV90" J. Bacteriol.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] S. Yamamoto, M. Nozaki, and Y. Ishimura (editors): "International Symposium on Oxygenases and Oxygen Activation" Yamada Science Foundation. 1-217 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K.Horiike,T.Kobayashi,T.Ishida and M.Nozaki: "οーNitorophenol as an active site probe for catechol 2,3ーdioxygenase." Yamada Conference XXVII International Symposium on Oxyenases and Oxygen Activation,edited S.Yamamoto,M.Nozaki,Y.Ishimura. 15-18 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] M.Koyama,T.Ishida,K.Horiide,M.Nozaki and M.Shimada: "Urate oxidse and copper content in the liver of macular mutant mouse,a model animal for human congenital copper deficiency,Menkes kinky hair disease." J.Nutr.Sci.Vitaminol.(1992)

    • Related Report
      1991 Annual Research Report
  • [Publications] H.Tojo,K.Horiike,T.Ishida,T.Kobayashi,M.Nozaki and M.Okamoto: "Analytical and microーpreparative highーperformance gel chromatography of proteins with a short column of porous silicaーbased aqueous gels:Estimation of molecular size rapid determination of ligand binding constant,and purification of Sーcarboxymethylated proteins for microsequecing." J.Chromatography. (1992)

    • Related Report
      1991 Annual Research Report
  • [Publications] H.Fujioka,K.Horiike,M.Takahashi,T.Ishida,M.Kinoshita and M.Nozaki: "Triphasic Vascular effects of sulfhydrylーcontaining compounds and Their oxidized forms on dog coronary arteries." Life Science. (1992)

    • Related Report
      1991 Annual Research Report
  • [Publications] Manzoor A.Bhat,T.Ishida K.Horiike,C.S.Vaidyanathan and M.Nozaki: "Purification of 3,5ーdichlorocatechol 1,2ーdioxygenase A nonheme iron dioxygenase and a key enzyme in the biodegradation of a herbicide 2,4ーdichlorophenoxyacetic acid (2,4ーD) from pseudomonas cepacia CSV90" J.Biol.Chem.

    • Related Report
      1991 Annual Research Report
  • [Publications] Manzoor A.Bhat,et al.: "Isolation and characterization of a new plasmid encoding genes for the degradation of 2,4ーdichlorophenoxyacetate (2,4ーD) from pseudomonas cepacia CSV90" J.Bacteriol.

    • Related Report
      1991 Annual Research Report
  • [Publications] S.Yamamoto,M.Nozaki,and Y.Ishimura (editors): "International Symposium on Oxygenases and Oxygen Activation" Yamada Science Foundation, 217 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] C.Nakai,K.Horiike,S.Kuramitsu,H.Kagamiyama and M.Nozaki: "Three isozymes of catechol 1,2-dioxygenase (pyrocatechase),αα,αβ,and ββ,from pseudomonas arvilla Cー1." J.Biol.Chem.205. 660-665 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] M.Nozaki,C.Nakai,K.Horiike,S.Kuramitsu,and H.Kagamiyama(edited by C.C.Reddy,G.A.Hamilton and K.M.Medyastha): "Three isozymes of catechol 1,2-dioxygenase (pyrocatechase) from Pseudomonas arvilla Cー1." Biological Oxidation Systems. 1. 405-416 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] 野崎 光洋,山本 尚三: "酵素による酸素化反応" 季刊化学総説. 7. 147-161 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] K.Horiike,T.Kobayashi T.Ishida and M.Nozaki(edited by S.Yamamoto,M.Nozaki,Y.Ishimura): "oーNitorophenol as an active site probe for catechol 2,3-dioxygenase." Yamada Conference XXVII International Symposium on Oxygenase and Oxygen Activation. 15-18 (1991)

    • Related Report
      1990 Annual Research Report
  • [Publications] 村松 正實・永井 克孝・本庶 佑(監訳),野崎 光洋(分担翻訳): "ストライヤ-生化学" バイオメディックス, 1092 (1991)

    • Related Report
      1990 Annual Research Report

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Published: 1991-04-01   Modified: 2016-04-21  

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