| Project/Area Number |
10670257
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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 | Okayama University |
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Principal Investigator |
YAMAMOTO Shigeo Okayama University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (40033229)
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| Co-Investigator(Kenkyū-buntansha) |
NARIMATSU Shizuo Okayama University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (20113037)
SHINODA Sumio Okayama University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (50029782)
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| Project Period (FY) |
1998 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2000: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1999: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1998: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | Pathogenic vibrio / Iron-reressible gene / Gene sequencing / Iron acquisition system / Pathogenic factor / Stress response / Iron transport / Molecular genetics / 遺伝子クローニング / 腸炎ビブリオ / 鉄輸送 / 外膜蛋白 / 鉄欠乏症 / 鉄レギュロン |
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| Research Abstract |
1. V.parahaemolyticus strains isolated from different sources were assayed for their ability to produce a siderophore, vibrioferrin (VF), under iron-limited conditions. VF production of clinical isolates was significantly higher than those for food and coastal isolates, suggesting that greater producitivity of VF by clinical isolates may be associated with a selective advantage for survival and proliferation under conditions of iron-limitation such as in the intestine.
2. V.parahaemolyticus strains carrying mutations in the fur gene were isolated by the Mn selection method to assess the function of Fur in connection with alterations in the coordinate expression of the siderophore VF and iron-repressive outer membrane proteins (IROMPs). Ten of 25 mutants constitutively produce VF and expressed at least two IROMPs. DNA sequencing of the fur genes identified 4 different point mutations causing amino acid change. Western blotting using anti-Fur antiserum revealed that the intracellular abun
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dance of the wild-type Fur was not significantly affected by the iron concentration in the growth medium. Fur regulon was identified for the first time in V.parahaemolyticus.
3. Many kinds of iron-regulated gene fragments containing a putative Fur box were isolated from pathogenic vibrios by their nucleotide sequencing. The full-length genes encoding three IROMPs in V.parahaemolyticus were cloned and analyzed. Also, the aerobactin operon and heme receptor were cloned and analyzed from V.mimicus. The genetic organization of the former was unusual in that it contains the genes responsible for the iron transport system (fhuBCD) in addition to the genes encoding biosynthesis enzymes (iucABCD) and an outer membrane receptor (iutA).
4. Iron-repressible sodA genes encoding MnSOD were cloned from three vibrio species, and the Fur boxes were identified in their promoters. These MnSODs were produced in response to iron-limitation.
5. One of the genes responsible for VF biogenesis was identified by insertion mutagenesis using a suicide vector. The insertion mutagensis was applied to the above-described V.parahaemolyticus genes to clarify their functions.
6. The genes encoding ATP-dependent protease were isolated from three vibrio species. It is of great interest to clarify whether the iron-regulated protease expression is associated with decomposition of Fur in response to iron starvation. Less
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