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Structure and Function of Histidinol phosphate Aminitrans ferase

Research Project

Project/Area Number 14580632
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Structural biochemistry
Research InstitutionOsaka Medical College

Principal Investigator

MIZUGUCHI Kiroyuki  Osaka Medical College, Faculty of Medicine, Research Associate, 医学部, 助手 (40247838)

Project Period (FY) 2002 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 2003: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2002: ¥1,200,000 (Direct Cost: ¥1,200,000)
KeywordsAminotransferase / Pyridoxal 5'-phosphate / Schiff base pK_a / Reaction mechanism / Strain / ヒスチジノールリン酸アミノ基転移酵素 / グルタミンアミノ基転移酵素 / ピリドキサルリン酸 / シッフ塩基 pK_a
Research Abstract

HPAT followed a Ping-Pong Bi Bi kinetic mechanism. The PLP-form HPAT showed pH dependent absorption and CD spectral changes with maxima at 340 nm at high pH and 420 nm at loW pH (pK_a=6.6). Absorption spectral changes of the glutarate-or 2-methylglutamate-bound wild type enzyme indicated that the Schiff base pK_a of the Michaelis complex and the external aldimine were increased. Absorption spectra of K214A mutant enzyme in the presence of 2-methylglutamate demonstrated that the intrinsic Schiff base pK_a of the external aldimine was much elevated. The pK_a value of the methylamine-substituted K214A mutant enzyme was increased from 6.6 to 10.6. Mutation of Asn157 to Ala increased the pK_a to 9.2. Data indicate that the strain of the Schiff base is the principal factor to decrease the pK_a in HPAT, and is crucial for the subsequent increase in the Schiff base pK_a during catalysis. It is suggested that the Schiff base strain mechanism is a common to the Subgroup I aminotransferases.

Report

(3 results)
  • 2003 Annual Research Report   Final Research Report Summary
  • 2002 Annual Research Report
  • Research Products

    (15 results)

All Other

All Publications (15 results)

  • [Publications] Hiroyuki Mizuguchi: "Characterization of histidinol phosphate aminotransferase from Eschericia coli"Biochim Biophys Acta.. 1647. 321-324 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hideyuki Hayashi: "Strain and catalysis in aspartate aminotransferase"Biochim.Biophys.Acta.. 1647. 103-109 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hideyuki Hayashi: "Conformational change in aspartate aminotransferase on substrate binding induces strain in the catalytic group and enhances catalysis"J.Biol.Chem.. 278. 9481-9488 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Akira Hosono: "Glutamine : phenylpyruvate Aminotransferase from an Extremely Thermophilic Bacterium, Thermus thermophilus HB8"J.Biochem.. 134. 843-851 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Rie Omi: "Crystal Structures of Threonine Synthase from Thermus thermophilus HB8:Conformational change, substrate recognition, and mechanism."J.Biol.Chem.. 278. 46035-46045 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Masaru Goto: "Crystal Structures of Glutamine : Phenylpyruvate Aminotransferase from Thermus thermophilus HB8. INDUCED FIT AND SUBSTRATE RECOGNITION^*"J.Biol.Chem.. 279. 16518-16525 (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Mizuguchi, H., Hayashi, H., Miyahara, I., Hirotsu, K., Kagamiyama, H.: "Characterization of histidinol phosphate aminotransferase from Escherichia coli."Biochim.Biophys.Acta. 1647. 321-324 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hayashi, H., Mizuguchi, H., Miyahara, I., Islam, M.M., Ikushiro, H., Nakajima, Y., Hirotsu, K., Kagamiyama, H.: "Strain and catalysis in aspartate aminotransferase."Biochim.Biophys.Acta.. 1647. 103-109 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hayashi, H., Mizuguchi, H., Miyahara, I., Nakajima, Y., Hirotsu, K., Kagamiyama, H.: "Conformational change in aspartate aminotransferase on substrate binding induces strain in the catalytic group and enhances catalysis."J.Biol.Chem.. 278. 9481-9488 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Omi, R., Goto, M., Miyahara, I., Mizuguchi, H., Hayashi, H., Kagamiyama, H., Hirotsu, K.: "Crystal Structures of Threonine Synthase from Thermus thermophilus HB8 : Conformational change, substrate recognition, and mechanism."J.Biol.Chem.. 278. 46035-46045 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hosono, A., Mizuguchi, H., Hayashi, H., Goto, M., Miyahara, I., Hirotsu, K., Kagamiyama, H.: "Glutamine : phenylpyruvate Aminotransferase from an Extremely Thermophilic Bacterium, Thermus thermophilus HB8"J.Biochem.. 134. 843-885

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Goto, M., Omi, R., Miyahara, I., Hosono, A., Mizuguchi, H., Hayashi, H., Kagamiyama., H., Hirotsu, K.: "Crystal Structures of Glutamine : phenylpyruvate Aminotransferase from Thermus thermophilus HB8 INDUCED FITAND SUBSTRATE RECOGNITION^*"J.Biol.Chem.. 279. 16518-16525 (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Hiroyuki Mizuguchi: "Characterization of histidinol phosphate aminotransferase from Escherichia"Biochim Biophys Acta.. 1647(1-2). 321-324 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Akira Hosono: "Glutamine : phenylpyruvate Aminotransferase from an Extremely Thermophilic Bacterium, Thermus thermophilus HB8"Journal of Biochemistry. 134(6). 843-851 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Hiroyuki Mizuguchi: "Characterization of histidinol phosphate aminotransferase from Eschericia coli"Biochimica et Biophysica Acta. (in press). (2003)

    • Related Report
      2002 Annual Research Report

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Published: 2002-04-01   Modified: 2016-04-21  

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