Grant-in-Aid for International Scientific Research.
|Section||Joint Research .|
|Research Institution||Osaka University|
TANIGUCHI Naoyuki Medical School, Osaka University, 医学部, 教授 (90002188)
SCHACHTER Ha トロント大学, 医学部, 教授
SIEST Gerard University of Nancy I Faculty of Sciences France, 薬学生物部, 教授
DEUTSCH Haro ウィスコンシン大学, 医学部, 名誉教授
ANDERSON Mar コーネル大学, 医学部, 助教授
近藤 宇史 長崎大学, 医学部, 教授 (00158908)
井原 義人 大阪大学, 医学部, 助手 (70263241)
藤井 順逸 大阪大学, 医学部, 助教授 (00222258)
CHOUVAEV Vla ナンシー第一大学, 医学部, 研究員
YAMAMOTO Hir シカゴ神経科学研究所, 主任研究員
DENNIS James トロント大学, 医学部, 教授
CHOUVAEV Vladimir V University of Nancy I Faculty of Medicine France
YAMAMOTO Hirotaka Chicago Institute of Neurosergery U.S.A.
DENNIS James W University of Toronto, Mesical Scienees Canada
DEUTSCH Harold F University of Wisconsin Medical Center U.S.A.
ANDERSON Mary E Cornell Medical Center U.S.A.
IHARA Yoshito Medical School, Osaka University
SCHACHTER Harry Medical Sciences, University of Toronto, Canada
FUJII Junichi Medical School, Osaka University
KONDO Takahito School of Medicine Nagasaki University
|Project Fiscal Year
1995 – 1996
Completed(Fiscal Year 1996)
|Budget Amount *help
¥10,600,000 (Direct Cost : ¥10,600,000)
Fiscal Year 1996 : ¥5,300,000 (Direct Cost : ¥5,300,000)
Fiscal Year 1995 : ¥5,300,000 (Direct Cost : ¥5,300,000)
|Keywords||diabetes / vascular smooth muscle / glycation / site-directed mutagenesis / apolipoprotein E / heparin binding EGF-like growth factor / gamma-glutamyl transpeptidase / 糖尿病 / 血管平滑筋 / 糖化反応 / 部位特異的変異 / アポリポプロテインE / ヘパリン結合性EGF様増殖因子 / γ-グルタミルトランスペプチダーゼ / r-グルタミルトランスペプチダーゼ / 糖鎖遺伝子 / γ-glutamyl transpeptidase / 糖タンパク質 / 糖転移酵素 / アポプロテインE|
gamma-Glutamyl transpeptidase, a heterodimeric glycoprotein anchored to the extra cellular surface of cell membrane, plays an important role in glutathione metabolism. It catalyzes the tansfer reaction of a gamma-glutamyl moiety from glutathione and related compounds to variety of amino acids and dipeptides. We have established tha expression system for gamma-glutamyl transpeptidase in vaculovirus/insect system and analyzed the active sites of the enzyme. We found the serine residues required for batalysis of the enzyme was identified by site-specific mutagenesis of the conserved serine residues on the basis of required alignment of the light subbing of human, rat pig and two bacterial enzymes. Recombinant human enzyme with replacements of these serine residues by Ala were expressed using a baculovirus-insect cell system. There results suggest that both Ser-45tl and Ser-452 are located at the position able to interact with the gamma-glutamyl group and participate in catalysis, probably
as nucleophiles or through stabilization of the transition state and the residues ware found to be well conserved in various animal and plant species. These works have been carried out in collaboration with Dr.Alton meister and Dr.Mary Anderson at the Cornell University Medical School in New York and we have published several papers in J.Biol. Chem and Proc.Natl.Acad.Sci.USA.
Apolipoprotein E (ApoE) undergoes non-enzymatic glycosylation (glycation) under hyperglycemic conditions such as diabetes and results in the decrease of total binding and affinity to heparin. In plasma, ApoE also undergoes glycation in the very low density lipoprotein fractions under normal and diabetic conditions. In diabetic patients however, glycation is dramatically increased up to 2-3 folds. Moreover, ApoE stimulates the activity of heparin binding EGF like growth factor (HB-EGF) which is a mitogen to vascular smooth muscle cells. These results indicate that both apo E and HB-EGF may play a critical role in the pathogenesis of atherogenic changes in the diabetes. These works have been done in collaboration with Drs.Gerard Siest, Maria Wellman and Vladimir Shuvaev at Nancy, France. Less