| Project/Area Number |
14570252
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Bacteriology (including Mycology)
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| Research Institution | Tokushima Bunri University |
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Principal Investigator |
YAMANAKA Hiroyasu Tokushima Bunri University, Pharmaceutical Sciences, Associate Professor, 薬学部, 助教授 (30202386)
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| Co-Investigator(Kenkyū-buntansha) |
OKAMOTO Keinosuke Okayama University, Pharmaceutical Sciences, Professor, 薬学部, 教授 (70131183)
竹治 美穂 徳島文理大学, 薬学部, 助手 (20320103)
野村 知彦 徳島文理大学, 薬学部, 助手 (00289315)
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| Project Period (FY) |
2002 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2004: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2003: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2002: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | Escherichia coli / Heat-stable enterotoxin / Outer membrane transport / TolC / Drug efflux / Processing |
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| Research Abstract |
Escherichia coil outer membrane protein TolC is involved in the export of chemically diverse molecules such as antibiotics, bile salts, detergents, and various organic solvents. TolC is also engaged in the extracellular secretion of protein toxins such as a-hemolysin and heat-stable enterotoxin (ST) in some pathogenic strains of E.coli. Therefore, we believe that the study on the function of TolC contributes to clarifying an extracellular transport process in E.coli and also provides useful information preventing infection from the pathogenic strains.
Our previous studies indicated that the leucine residues at positions 3 (Leu-3) and 412 (Leu-412) are essential for the transport activity of TolC. In this study, we examined the transport activities of TolC homologues derived from Salmonella typhimurium and Vibrio parahaemolyticus in E.coli and identified a functional domain of E.coll TolC.
1.Both S.typhimurium and V.parahaemolyticus tolC genes were cloned into pET-STI plasmid. The primary structures of S.typhimurium TolC (S-TolC) and V.parahaemolyticus TolC (VP-TolC) were 89.5% and 43.3% homologous to that of E.coli TolC (EC-TolC), respectively.
2.The secretory efficiency of ST to the exterior was largely (50%) decreased in the VP-TolC- expressing E.coli, but in the S-TolC-expressing E.coli was not. This result indicates that VP-TolC can not fully function in E.coli cells. We think that the reason is ascribed to the existence of low homologous regions in VP-TolC to EC-TolC.
3.We further examined the transport activities using several E.coli transformants producing various EC/VP- chimera TolCs. The results suggest that the region containing amino acid residues from 198 to 214 of EC- TolC is important to express the transport activity in E.coli cells. We think that the region must be a functional domain which changes the TolC channel from a closed state to an open state. Further studies are currently in progress in our laboratory.
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