| Project/Area Number |
04680160
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
物質生物化学
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| Research Institution | Osaka University |
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Principal Investigator |
OZAKI Hiroshi Osaka University, Institute for Protein Research, Instructor, 蛋白質研究所, 助手 (90116012)
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| Project Period (FY) |
1992 – 1993
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| Project Status |
Completed (Fiscal Year 1993)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1993: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1992: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | Heat-stable / Enterotoxin / ST-receptor / Fluorophor / Purification / Membrane protein |
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| Research Abstract |
Heat-stable enterotoxins (STs) are peptides of 18 or 19 amino acid residues produced by enterotoxigenic Escherichia coli and are known to cause acute diarrhea in men and animals. The biological action of ST is initiated by the binding to its receptor protein on the membrane of intestinal epithelial cell. The formation of a ST-receptor complex leads to stimulation of guanylate cyclase and is followed by increase in GMP concentration in the cells and fluid secretion from the cells. This study is planned to identify the binding part of the receptor protein in its binding to ST, purify and crystallize the complex j of ST with its receptor protein for its X-ray crystallographic analysis in order to elucidate the co-recogniton mechanism between ST and its receptor protein which is the first step of signal transduction by ST via the receptor protein on the membrane of cells. To accomplish this purpose, I synthesized a variety of ST analogues were labeled with the fluorophore and cific binding abilties of these ST analogues were examined by means of chromatographic method. Among these, there were two ST analogues with the specific binding ability to the receptor protein corresponding to that of the native ST.One of these analogues was used to form the complex with the receptor protein on the membrane prepared from rat intestine. After solubilization experimental conditions to purify the complex were examined and the conbination of several kinds of HPLC lead to obtain a partially purified complex with fluorescence to get the knowledge about the binding part of receptor protein to ST, the purification of the protein crosslinked with ST labeled with the fluorophore is in progress.
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