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

Role of Conserved Proline Residues in Conformation, Function, and Stability of a Protein

Research Project

Project/Area Number 02680134
Research Category

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

Allocation TypeSingle-year Grants
Research Field 物質生物化学
Research InstitutionOsaka University

Principal Investigator

YUTANI Katsuhide  Inst. Protein Res., Osaka Univ., Assoc. Prof., たんぱく質研究所, 助教授 (90089889)

Project Period (FY) 1990 – 1991
Project Status Completed (Fiscal Year 1991)
Budget Amount *help
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1991: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1990: ¥900,000 (Direct Cost: ¥900,000)
KeywordsTryptophan Synthase / Scanning Calorimetry / Isothermal Calorimetry / Conformational Stability / CD / Proline / トリプトファン合成酵素 / 蛋白質の変性 / 蛋白質の構造形成 / 変異蛋白質
Research Abstract

To study the role of Pro residues in the conformation, stability, and function of a protein, nine mutant alpha-subunits of tryptophan synthase from Escherichia coli, in which Ala or Gly was substituted for each of six conserved Pro residues(positions 28, 57, 62, 96, 132 and 207)in 10 microorganisms, were constructed.
1) The far-UV CD spectra of five mutant alpha-subunits with Ala in place of Pro, the exception being the mutant at position 207(P207A), were identical to the spectrum of the wild-type protein. CD values in the far-UV region were less negative for P207A, indicating that the Pro residue at position 207 plays a role in maintaining the intact structure of the alpha-subunit.
2)Scanning calorimetric measurements(DASM4)showed that the stability of each mutant protein relative to that of the wild-type was about the-same for P57A, less for P62A and P132A, and markedly decreased for P96A and P207A ; which are substituted at less mobile positions.
3)To understand how the alpha and beta_2 subunits of tryptophan synthase interact to form an alpha_2beta_2 complex and undergo mutual activation, we have investigated isothermal calorimetric titrations (Omega) of wild type beta_2 subunit with wild type alpha subunit and a mutant alpha subunit containing a substitution of Gly for Pro at position 132 show that both the affinity and the exothermic association enthalpy are greatly reduced in the mutant alpha subunit although the stoichiometry of association is unchanged. We conclude that Pro 132 plays a critical role in subunit interaction and in mutual subunit activation.

Report

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

    (23 results)

All Other

All Publications (23 results)

  • [Publications] K.Yutani,S.Hayashi,Y.Sugisaki,& K.Ogasahara: "Role of Conserved Proline Residues in Stabilizing Tryptophan Synthase α-Subunit:Analyzed by Mutants with Alanine or Glycine." Proteins,. 9. 90-98 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] T.Hering,K.Yutani,Y.Taniyama,& M.Kikuchi: "Effect of Proline Mutations on the Unfolding and Refolding of Human Lysozyme:The slow Refolding Kinetic Phase Does not Results from Proline Cis-Trans Isomerization." Biochemistry. 30. 9882-9891 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K.Ogasahara,K.Hiraga,W.Ito,E.W.Miles,& K.Yutani: "Origin of the Mutual Activation of the α and β_2 Subunits in the α_2β_2 Complex of Tryptophan Synthase:Effect of Alanine or Glycine Substitutions at Proline Residues in the α Subunit" 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] Y.Taniyama,K.Ogasahara,K.Yutani,& M.Kikuchi: "Folding Mechanism of Mutant Human Lysozyme C77/95A with Increased Secretion Efficiency in Yeast." 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] 油谷 克英: "蛋白質立体構造の構築原理は「アミノ酸置換の手法」でどこまで解明できたか" 「生物物理」. 32. 27-32 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] S. Sawada, A. Hideo, K. Ogasahara, and K. Yutani.: "Assignment of Tyrosine Resonances in Proton NMR Spectrum of Tryptophan Synthase alpha-Subunit. Monitering Conformational Changes due to Substitutions at Position 49." Eur. J. Biochem.189. 667-673 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Y. Sugisaki, K. Ogasahara, E. W. Miles, and K. Yutani: "Scanning Calorimetric Study of Tryptophan Synthase alpha-Subunit from Escherichia Coli, Salmonella Typhimurium, and Interspecies Hybrid." Thermochimica Acta.163. 117-122 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K. Yutani, S. Hayashi, Y. Sugisaki, and K. Ogasahara.: "Role of Conserved Proline Residues in Stabilizing Tryptophan Synthase alpha-Subunit : Analyzed by Mutants with Alanine or Glycine." Proteins. 9. 90-98 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] T. Hering, K. Yutani, Y. Taniyama, and M. Kikuchi: "Effect of Proline Mutations on the Unfolding and Refolding of Human Lysozyme : The slow Refolding Kinetic Phase Does not Results from Proline Cis-Trans Isomerization." Biochemistry. 30. 9882-9891 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K. Ogasahara, K. Hiraga, W. Ito, E. W. Miles, and Katsuhide Yutani: "Origin of the Mutual Activation of the alpha and beta_2 Subunits in the alpha2beta_2 Complex of Tryptophan Synthase : Effect of Alanine or Glycine Substitutions at Proline Residues in the alpha Subunit" J. Biol. Chem.267. 5222-5228 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] Y. Taniyama, K. Ogasahara, K. Yutani, and M. Kikuchi: "Folding Mechanism of Mutant Human Lysozyme C77/95A with Increased Secretion Efficiency in Yeast." J Biol. Chem.267. 4619-4624 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] K.Yutani,S.Hayashi,Y.Sugisaki,& K.Ogasahara: "Role of Conserved Proline Residues in Stabilizing Tryptophan Synthase αーSubunit:Analyzed by Mutants with Alanine or Glycine." Proteins. 9. 90-98 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] T.Hering,K.Yutani,Y.Taniyama,& M.Kikuchi: "Effect of Proline Mutations on the Unfolding and Refolding of Human Lysozyme:The slow Refolding Kinetic Phase Does not Results from Proline CisーTrans Isomerization." Biochemistry. 30. 9882-9891 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] K.Ogasahara,K.Hiraga,W.Ito,E.W.Miles,& K.Yutani: "Origin of the Mutual Activation of the α and β_2 Subunits in the α_2β_2 Complex of Tryptophan Synthase:Effect of Alanine or Glycine Substitutions at Proline Residues in the α Subunit" J.Biol.Chem.

    • Related Report
      1991 Annual Research Report
  • [Publications] Y.Taniyama,K.Ogasahara,K.Yutani,& M.KiKuchi: "Folding Mechanism of Mutant Human Lysozyme C77/95A with Increased Secretion Efficiency in Yeast." J.Biol.Chem.

    • Related Report
      1991 Annual Research Report
  • [Publications] 油谷 克英: "蛋白質立体構造の構築原理は「アミノ酸置換の手法」でどこまで解明できたか" 「生物物理」. 32. 27-32 (1992)

    • Related Report
      1991 Annual Research Report
  • [Publications] 油谷 克英,中村 春木: "蛋白質工学" 朝倉書店、東京, 208 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Sugisaki,Y.,Ogasahara,K.,Miles,E.W., & Yutani,K.: "Scanning Calorimetric study of Tryptophan Synthase αーSubunit from E.coli,S.typhimurium and an Interspecies Hybrid" Thermochimica Acta. 163. 117-122 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] Yutani,K.,Hayashi,S.,Sugisaki,Y.,& Ogasahara,K.: "Role of Conserved Proline Residues in Stabilizing Tryptophan Synthse αーSubunit:Analysis by Mutants with Alanine or Glycine" PROTEINS Structure,Function,and Genetics. 9. 90-98 (1991)

    • Related Report
      1990 Annual Research Report
  • [Publications] Go,M.,Tomoda,T.,Honda,M.,& Yutani,K.: "Domain and Module Structure in α/β Barrel of Tryptophan Synthase α Subunit" PROTEINS Structure,Function,and Genetics.

    • Related Report
      1990 Annual Research Report
  • [Publications] 小笠原 京子,油谷 克英: "トリプトファン合成酵素の構造と機能" 蛋白質核酸酵素. 35. 201-211 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] Yutani,K.& Ogasahara,K.(Edited by Hatano,M.): "Protein Structural Analysis,Folding and Design" JapanScientific Societies Press Elsevier, 237 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] 油谷 克英(分担執筆): "新生化学実験講座第1巻タンパク質III高次構造" 東京化学同人(日本生化学会編), 436 (1990)

    • Related Report
      1990 Annual Research Report

URL: 

Published: 1990-04-01   Modified: 2016-04-21  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi