2000 Fiscal Year Final Research Report Summary
Identification and purification of protective antigen on Vibrio cholerae O1 infection
| Project/Area Number |
10670263
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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 | University of the Ryukyus |
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Principal Investigator |
NAKASONE Noboru Faculty of Medicine, University of the Ryukyus, Assistant Professor, 医学部, 助手 (80175497)
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| Co-Investigator(Kenkyū-buntansha) |
HONMA Yasuko Fujita Health University School of Medicine Associate Professor, 医学部, 助教授 (90253955)
IWANAGA Masaaki Faculty of Medicine, University of the Ryukyus, Professor, 医学部, 教授 (00112384)
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| Project Period (FY) |
1998 – 2000
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| Keywords | Vibrio cholerae O1 / colonization factor / hemagglutinin / OmpU / in vivo |
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| Research Abstract |
The outermembrane protein OmpU, Fucose sensitive hemagglutinin (FSHA) and in vivo expressed outermembrane protein P7R of Vibrio cholerae were investigated to identify colonization factor (s) of V.cholerae.
OmpU was purified and characterized. The purified OmpU adhered to the rabbit intestinal epithelium, however, the adhesion was not inhibited by treating the organisms with anti-OmpU and treating the intestine with purified OmpU.The antibody did not agglutinate the organisms, suggesting OmpU is not surface exposed. Therefore, OmpU is not supposed to function as a colonization factor.
FSHA could not be purified by our methods, therefore, genetic approach was made. Transposon (Tn5) was randomly inserted to the chromosome of V.cholerae O1 strains 86B3, and hemagglutinin negative mutant was screened. Now we are on the process of identifying the position of hemagglutination gene.
P7R is outermembrane protein which express in vivo. The molecular weight of P7R was estimated to be about 17 kDa by SDS-polyacrylamide get electrophoresis. N-terminal amino acid sequence of P7R were AEILKSDGTVDFYEQLR, and showed a high degree of homology with V.cholerae OmpT.Antisera of cholerae patients were reacted to P7R.However, the adhesion was not inhibited by treating the organisms with anti-P7R Fab fractions. The antibody of P7R did not agglutinate the organisms grown in vitro. Therefore, the adhesion inhibition test should do in vivo experimental conditions. The experiment are now under processing.
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