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Protein Engineering for New Functional Hemoproteins Based on Module Substitution

Research Project

Project/Area Number 07409003
Research Category

Grant-in-Aid for Scientific Research (A)

Allocation TypeSingle-year Grants
Section一般
Research Field 広領域
Research InstitutionKYOTO UNIVERSITY

Principal Investigator

MORISHIMA Isao  Kyoto University, Graduate School of Engineering, Professor, 工学研究科, 教授 (50026093)

Co-Investigator(Kenkyū-buntansha) TAKAHASHI Satoshi  Kyoto University, Graduate School of Engineering, research Associate, 工学研究科, 助手 (30283641)
ISHIMORI Koichiro  Kyoto University, Graduate School of Engineering, Associate Professor, 工学研究科, 助教授 (20192487)
Project Period (FY) 1995 – 1997
Project Status Completed (Fiscal Year 1997)
Budget Amount *help
¥30,700,000 (Direct Cost: ¥30,700,000)
Fiscal Year 1997: ¥4,800,000 (Direct Cost: ¥4,800,000)
Fiscal Year 1996: ¥5,900,000 (Direct Cost: ¥5,900,000)
Fiscal Year 1995: ¥20,000,000 (Direct Cost: ¥20,000,000)
KeywordsModule / Exon Shaffling / Globin / エクソンジャフリング
Research Abstract

The primary results in this research project are as followa :
(1)Module Substitution in Globins and Peroxidases. The structural and functional properties of the module-substituted globins and peroxidases have been characterized. Most of the module-substituted proteins exhibited highly destabilized protein structure, probably due to the missing or severe perturbation in the key interactions between the modules. The destabilized protein structure also interfered the formation of the specific dimers or tetramers in globins and enzymatic activity in peroxidases. It can be, therefore, concluded that the module-module interactions are also essential to design new stable and functional proteins as well as the module substitution.
(2)Identification of New Module Structure in Globins. The amino acid substituteions based on the structural unit of wchic boundaries are located at the middle of the conventional modules have revealed that the module previously proposed can be divided into new and smaller structural units. In this research project, I focused upon one of the newly identified structural units, which includes the iron-liganded histidine and about 20 amino acid residues in the proximal site, "sub-module m6". The substitution of the sub-module m6 in globins drastically affect the electronic state of heme and configuration of the iron-liganded histidine, indicating that the sub-module m6 would be a "heme binding modules" respondible for the heme binding structure.

Report

(4 results)
  • 1997 Annual Research Report   Final Research Report Summary
  • 1996 Annual Research Report
  • 1995 Annual Research Report
  • Research Products

    (19 results)

All Other

All Publications (19 results)

  • [Publications] Matsui, T., Nagano, S., Ishimori, K, et al.: "Preparation and Reaction of Myoglobin Mutants Bearing Both Proximal Cysteine Ligand and Hydrophobic Distal Cavity : Protein Models for the Active Site of P-450" Biochemistry. 35. 13118-13124 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Nagano, S., Tanaka, M., Ishimori, K., et al.: "The Catalytic Roles of the Distal Site Asparagine-Histidine Couple in Peroxidases" Biochemistry. 35. 14251-14258 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Tanaka, M., Ishimori, K., Morishima, I.: "The Distal Glutamic Acid As Acid-Base Catalyst in the Distal Site of Horseradish Peroxidase" Biochem.Biophys.Res.Commun. 227. 393-399 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Wakasugi, K., Ishimori, K., Morishima, I.: ""Module"-substituted Globins : Artificial Exon Shuffling Among Myoglobin,Hemoglobin α-and β-subunits" Biophys.Chem.68. 265-273 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Inaba, K., Wakasugi, K., Ishimori, K., Konno, T., Kataoka, M., Morishima, I.: "Structural and Functional Roles of Modules in Hemoglobin.-Substitution of Module M4 in Hemoglobin Subunits-" J.Biol.Chem.272. 30054-30060 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Inaba, K., Ishimori, K., Imai, K., Morishima, I.: "Ststructural and Functional Effects of Pseudo-module Substitution in hemoglobin Subunits : New Structural and Functional Units in Globin Structure" J.Biol.Chem.273(印刷中). (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Matsui, T., Nagano, S., Ishimori, K., et al.: "Preparation and Reaction of Myoglobin Mutants Bearing Both Proximal Cysteine Ligand and Hydrophobic Distal Cavity : Protein Models for the Active Site of P-450" Biochemistry. 35. 13118-13124 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Nagano, S., Tanaka, M,Ishimori, K., et al.: "The Catalytic Roles of the Distal Site Aspargine-Histidine Couple in Peroxidases" Biochemistry. 35. 14251-14258 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Tanaka, M., Ishimori, K., Morishima, I.: "The Distal Glutamic Acid As Acid-Base Catalyst in the DIstal Site of Horseradish Peroxidese" Biochem.Biophys.Res.Commun.227. 393-399 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Wakasugi, K., Ishimori, K., Morishima, I.: ""Module"-substituted Globins : Artificial Exon Shuffling Among Myoglobin, Hemoglobin alpha-and beta-subunits" Biophys.Chem.68. 265-273 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Inaba, K., Wakasugi, K., Ishimori, K., et al.: "Structural and Functional Roles of Modules in Hemoglobin. -Substitution of Module M4 in Hemoglibin Subunits-" J.Biol.Chem. 272. 30054-30060 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Inaba, K., Ishimori, K., Imai, K., Morishima, I.: "Structural and Functional Effects of Pseudo-module Substitution in hemoglobin Subunits : New Structural and Functional Units in Globin Structure" J.Biol.Chem. 272 (in press). (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Wakasugi,K., Ishimori,K., Morishima,I.: ""Module"-substituted Globins : Artificial Exon Shuffling Among Myoglobin,Hemoglobin α-and β-subunits" Biophys. Chem.68. 265-273 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Inada,K., Wakasugi,K., Ishimori,K., Konno,T., Kataoka,M., Morishima,I.: "Structural and Functional Roles of Modules in Hemoglobin. -Substitution of Module M4 in Hemoglobin Subunits-" J.Biol. Chem.272. 30054-30060 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Inada,K., Ishimori,K., Imai,K., Morishima,I.: "Structural and Functional Effects of Pseudo-module Substitution in hemoglobin Subunits : New Structural and Functional Units in Globin Structure" J.Biol.Chem.273(印刷中). (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Matsui,T: "Preparation and Reaction of Myoglobin Mutants Bearing Both Proximal Cysteine Ligand and Hydrophobic Distal Cavity : Protein Models for the Active Site of P-450" Biochemistry. 35. 13118-13124 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Nagano,S: "The Catalytic Roles of the Distal Site Asparagine-Histidine Couple in Peroxidases" Biochemistry. 35. 14251-14258 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Tanaka,M: "The Distal Glutamic Acid As Acid-Base Catalyst in the Distal Site of Horseradish Peroxidase" Biochem.Biophys.Res.Commun. 227. 393-399 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Morishima, I: "NMR Studies on Recombinant Cytochrome P450cam Mutants" Biochimie. (印刷中). (1996)

    • Related Report
      1995 Annual Research Report

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Published: 1995-04-01   Modified: 2016-04-21  

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