| Project/Area Number |
07044263
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Institution | Osaka University |
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Principal Investigator |
TANIGUCHI Naoyuki Medical School, Osaka University, 医学部, 教授 (90002188)
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| Co-Investigator(Kenkyū-buntansha) |
CHOUVAEV Vladimir V University of Nancy I Faculty of Medicine France, 医学部, 研究員
YAMAMOTO Hirotaka Chicago Institute of Neurosergery U.S.A., 主任研究員
DENNIS James W University of Toronto, Mesical Scienees Canada, 医学部, 教授
DEUTSCH Harold F University of Wisconsin Medical Center U.S.A., 医学部, 名誉教授
ANDERSON Mary E Cornell Medical Center U.S.A., 医学部, 助教授
SIEST Gerard University of Nancy I Faculty of Sciences France, 薬学生物部, 教授
SCHACHTER Harry Medical Sciences, University of Toronto, Canada, 医学部, 教授
IHARA Yoshito Medical School, Osaka University, 医学部, 助手 (70263241)
KONDO Takahito School of Medicine Nagasaki University, 医学部, 教授 (00158908)
FUJII Junichi Medical School, Osaka University, 医学部, 助教授 (00222258)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| 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)
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| Keywords | diabetes / vascular smooth muscle / glycation / site-directed mutagenesis / apolipoprotein E / heparin binding EGF-like growth factor / gamma-glutamyl transpeptidase / 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 |
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| Research Abstract |
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
… More
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
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