Analysis of the P.gingivalis hemagglutinin and the exploitation the replacement therapy
| Project/Area Number |
10671780
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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 |
病態科学系歯学(含放射線系歯学)
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| Research Institution | Nihon University |
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Principal Investigator |
YASUKO Shibata Nihon University School of Dentistry at Matsudo, Instructor, 松戸歯学部, 助手 (90133438)
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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 |
¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 2000: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1999: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1998: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | Periodontal desiase / Porphyromonas gingivalis / Hemagglutinin / Adherence / Specific anibody / 定着 / P.gingivalis / hemagglutinin / 定着阻止 / 抗原エピトープ |
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| Research Abstract |
Porphyromonas gingivalis is a gram-negative anaerobic bacterial species implicated as an important pathogen in the development of adult periodontitis. In our studies of P.gingivalis and ways to protect from periodontal disease, we have prepared the monoclonal antibody (MAb-Pg-vc) which is capable of inhibiting the hemagglutinating activity of P.gingivalis. In order to clarify the antigenically related hemagglutinating domains, we attempted to determine the minimum motifs responsible for P.gingivalis hemagglutinin. Initially, the 9 kb-EcoRI fragment encoding 130 kDa protein was cloned from the P.gingivalis chromosome using MAb-Pg-vc. Western blot analysis of nested deletion clones, the competition experiments using synthetic peptides, and the binding assay of the phage-displayed peptides using the MAb-Pg-vc allowed us to identify the minimum motifs, PVQNLT.Furthermore, the presence of multi-gene family coding for this epitope was confirmed via Southern blot analysis and PCR using the pr
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imers complementary to the domain corresponding to this epitope. It is suggested that the gene families specifying this short motif found in P.gingivalis chromosome encode many hemagglutinin and/or hemagglutinin-related protease molecules. This information might provide useful tools to establish passive immunization system to prevent periodontal disease.
Furthermore, to obtain a recombinant antibody, cDNAs coding for the variable domains of the L and H chains of MAb-Pg-vc were cloned using PCR techniques, and a plasmid specifying a single-chain variable fragment (ScFv) was constructed. Following transformation of Escherichia coli host, a recombinant ScFv protein was successfully expressed. This protein possessed exactly the same immunological properties as that of parental murine monoclonal antibody and the ScFv antibody inhibited the P.gingivalis vesicles-associated hemagglutinating activity. Since the expression system used in this study can readily provide large quantities of single chain recombinant antibody, it may be a-useful in developing a therapeutic agent for passive immunization against periodontal diseases. Next, we then attempted to produce a scFv antibody, which inhibited hemagglutination using the Bacillus brevis protein-producing system.
Furthermore, it was very interesting that the similarity was shown in the short motif (PVQNLT) associated with the hemagglutination of P.gingivalis and the top of the hemagglutinin molecule of Influenzae virus. So, we attempted to concern the cross reaction between the anti-hemagglutinating antibodies against the P.gingivalis and Influenzae virus. Less
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Report
(4 results)
Research Products
(9 results)
