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Construction of a soluble "glycochain-recognition protein" structure and regulation of embrionic/cellular mechanism using the protein apparatus

Research Project

Project/Area Number 09660359
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Applied molecular and cellular biology
Research InstitutionPrefecture University of Kyoto

Principal Investigator

OHNISHI Masatake  Prefecture University of Kyoto, Department of Biological Resources Chemistry, Professor, 農学部, 教授 (90026576)

Project Period (FY) 1997 – 1999
Project Status Completed (Fiscal Year 1999)
Budget Amount *help
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 1999: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1998: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1997: ¥2,400,000 (Direct Cost: ¥2,400,000)
Keywordsβ-galactosidase / β-glucocerebrosidase / Yeast / Japanese cattle / taste sense receptor / sulfite dehydrogenase / Thiobacillus thiooxidans / β-1, 3-exo-glucanase / β-ガラクトシダーゼ / ウシ脳 / ウシ舌 / 細胞膜装置 / β-グラクトシダーゼ / 配糖体 / 標的 蛋白質 / 膜酵素 / ベシクル / 甘味物質受容器 / 高粘度環境下酵素反応
Research Abstract

(1) β-Glycosidases from yeast (Kluyvelomyces lactis, Saccharomyces cerevisiae) : β-Galactosidases(GAL) and β-1, 3-exo-glucanase(LAM) were purified to be a single protein from K. lactis and S. cerevisiae, respectively, and were found to strongly catalyze their trasglycosilation reactions as follows ; GAL produces alkyl galactosides with alchols and galactosyl glyceride with glycerol. LAM produces laminari-oligosaccharides with laminari-biose and -triose. (2) β-Glucocerebrosidase(GCase) from cattle brain : GCase was purified in a soluble state(S) to analyze its enzymatic properties, and was immobilized on a carrier (decyl sepharose) to make a "glycochain-rcognition" structure. (3) Taste sense receptor protein from cattle tongue : Two receptor proteins, MW 58,000 and 64,000 were purified and treated with protease to have peptide fractions. Two major peptides each were obtained and analyzed their sequences to take a genetic operation for large scale production of these proteins. (4) Fish (Oryzia Latipes) germ cells : Freezing of sperm cells and egg cell culture were succeeded for around 4 days. Mechanism on reception of the fertilization signal (sperm cells) to the receptor on egg cells was investigated using the biological microscope obtained in this research project. (5) Sulfite dehydrogenase from Thibacillus thiooxidans : The microbe was employed for making of a high sensitive and automatical sulfite-analyzer as a sensor. Thus enzymatic bases (sulfite dehydrogenase-catalyzed reaction) were nedded to investigate about the microbial state using the kinetic procedures. Based on the subcellular fractionation, the enzyme was confirmed to be a membrane-bound state of organelles. It is very possible to constrast a "membrane-bound structure" for the analytical apparatus.

Report

(4 results)
  • 1999 Annual Research Report   Final Research Report Summary
  • 1998 Annual Research Report
  • 1997 Annual Research Report
  • Research Products

    (28 results)

All Other

All Publications (28 results)

  • [Publications] Ohnishi, M.: "An attempt to evaluate the subsite structure of cycloamylose glucanotransferase from Bacillus stearothermophilus"Starch. 49(9). 360-363 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Yazaki, T.: "Subsite structure of β-glucosidase from Aspergillus niger, Evaluated by steady-state kinetics with cell-ollgosaccharides as substrates"Carbohydrate Research. 298(1). 51-57 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Yazaki, T.: "Molecular mechanism on Aspergillus niger β-glucosidase-catalyzed reactions"Oyo Toshitsu-kagaku. 44(4). 523-529 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Yazaki, T,: "Stopped-flow kinetic studies on the binding of glucono-δ-lactone to- β-glucosidase from asporgillus niger"Oyo Toshitsu-kagaku. 45(1). 41-44 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Ohnishi, M.: "Characterization of the subsite structure of β-glucosidase from Aspergillus niger an aspect on the mechanism of carbohydrate ecognition"Carbohydr. Res.. 308(1). 201-205 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Matsumoto, T: "Kinetic study of sulfite dehydrogenase-catalyzed reaction in the subcellular state of Thiobacillus thiooxidans"J. Applied Glycoscience. 46(4)(in press). (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] 大西 正健: "酵素の科学"学芸出版センター. 222 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] 石嶌 純男: "細胞高分子学"学会出版センター. 161 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1999 Final Research Report Summary
  • [Publications] Matsumoto, T.: "A microbial biosensor for high sensitive and automatic detenmination of sulfite in foods and food materials"J. Applied Glycoscience. 46(3)in press. (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Matsumoto, T.: "Kinetic study of the sulfite dehydrogenase-catalyzed reaction in the subcellular state of Thiobaclllus thiooxidans"J. Applied Glycoscience. 46(4)in press. (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] 島田 睦: "ウシ脳グルコセレブロシダーゼに関する研究(1)-採取,精製,分子的特性の検討-"日本農芸化学会誌. 73. 26 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 藤村 裕子: "酵母Kluyveromyces lactis β-galactosidaseの転移反応"日本農芸化学会誌. 73. 33 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 松本 隆志: "食品中残存亜硫酸の高感度自動測定バイオセンサー装置、第1報硫黄酸化細菌の菌体による触媒反応の基盤解析"J. Applied Glycoscience. 46(3). 355 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 高橋 克夫: "グルコアミラーゼ反応におけるUrea効果、第1報、リガンド取り込み反応の速度論的解析"J. Applied Glycoscience. 46(3). 372 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 石嶌 純男: "細胞高分子学"学会出版センター. 161 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Ohnishi, M.: "Glycoenzymes"学会出版センター. 250 (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yazaki,T.: "Stopped-flow kinetic studies on the binding of glucono-δ-lactone to β-glucosidase from Aspergillus niger." Oyo Toshitus-kagaku. 44-1. 41-44 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Ohnishi,M.: "Characterization of the subsite structure of β-glucosidase from Aspergillus niger.an aspect on the mechanism of carbohydrate recognition." Carbohydrate Research. 308・1. 201-205 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 島田 睦: "Ureaによるグルコアミラーゼの構造変化と基質マルトース取り込み反応に及ぼす影響" 日本農芸化学会誌. 72(臨時増刊). 134- (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 矢崎 晃隆: "β-Glucosidaseのサブサイト構造とそのキャラクタリゼーション" 応用糖質科学. 45・2. 163-168 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 藤村 祐子: "酵母Kluyveromyces lactis β-galactosidaseに関する研究-精製及び分子的特性の検討-" 応用糖質科学. 45・3. 323- (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 島田 睦: "基質マルトオリゴ糖のグルコアミラーゼへの取り込み-細胞質酵素機能解析のモデル系に向けて-" 生化学. 70・8. 825- (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Ohnishi,M.: "An attempt to evaluate the subsite structure of cycloamyloseglucanotransferase from Bacillus stearothermophilus." Starch. 49・9. 360-363 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Yazaki,T.: "Subsite structure of β-Glucosidase from Aspergillus niger. Evaluated by steady-state kinetics with cello-oligosaccharides as substrates." Carbohydrate Research. 298・1. 51-57 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Yazaki,T.: "Molecular mechanism on Aspergillus niger β-glucosidase-catalyzed reactions." Oyo Toshitsu-kagaku. 44・4. 523-529 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Yazaki,T.: "Stopped-flow kinetic studies on the binding of glucono-δ-lactone to β-glucosidase from Aspergillus niger." Oyo Toshitsu-kagaku. 45・1. 41-44 (1998)

    • Related Report
      1997 Annual Research Report
  • [Publications] Ohnishi,M.: "Characterization of the subsite structure of β-glucosidase from Aspergillus niger. An aspect on the mechanism of carbohydrate recognition." Carbohydrate Research. (印刷中). (1998)

    • Related Report
      1997 Annual Research Report
  • [Publications] 島田 睦: "グルコアミラーゼ機能に及ぼすUreaの効果" 応用糖質科学. 44・3. 437 (1997)

    • Related Report
      1997 Annual Research Report

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Published: 1997-04-01   Modified: 2016-04-21  

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