| Project/Area Number |
12557188
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
矯正・小児・社会系歯学
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| Research Institution | Kagoshima University |
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Principal Investigator |
ITO Hiro-o Kagoshima University, Dental School, Associate Professor, 歯学部, 助教授 (40213079)
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| Co-Investigator(Kenkyū-buntansha) |
NAKASHIMA Toshihiro Chemo-Sero Therapeutic Research Institute, Research Science Manager, 主任研究員
SOUTOME Sakiko Kagoshima University, Dental Hospital, Research Associate, 歯学部附属病院, 助手 (20325799)
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| Project Period (FY) |
2000 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥12,200,000 (Direct Cost: ¥12,200,000)
Fiscal Year 2002: ¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 2001: ¥2,500,000 (Direct Cost: ¥2,500,000)
Fiscal Year 2000: ¥6,800,000 (Direct Cost: ¥6,800,000)
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| Keywords | Periodontal diseases / Adult periodontitis / Porphyromonas gingivalis / Monoclonal Antibodies / Vaccine / Fimbriae / Immunity / Phage-displayed peptide library / Porphyromonas gingivalis / ファージディスプレイ・ペプチドライブラリー / ファージ・ディスプレイ・ペプチドライプラリー |
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| Research Abstract |
Porphyromonas gingivalis is an important periodontal pathogen, and its fimbriae mediate the adherence of this bacterium to the host tissues. The aim of this study was to verify the possibility to generate a chemically synthesized vaccine which induces antibodies capable of inhibiting the ability of P. gingivalis to adhere host tissues. First, a panel of monoclonal antibodies (mAb) reacting to P. gingivalis fimbriae were established. All these mAb recognized higher order structure of fimbriae but not the primary or secondary structures. Next, a phage-displayed peptide library was constructed which contained an insertion of 18mer random peptide into the gp3 gene of M13 bacteriophage. Screening was performed according to the reactivity with one of the mAb, and a sequence probably mimicking the higher order structure of P. gingivalis fimbriae was identified. This phage clone was prepared and purified in a large quantity, and mice were immunized with the phage particles. The immunized mice
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produced serum antibodies specific for the inserted sequence and for native P. gingivalis fimbriae. To stabilize the conformational epitope of the selected peptide, the peptide was made cyclic on the phage by genetic engineering. As it was impossible to chemically synthesize the peptide, some amino acid residues in the sequence which were thought to be not important for the antigenicity were deleted or replaced, and a soluble cyclic peptide was chemically synthesized. However, these attempts did not improve the antigenicity of the peptide. It has been reported the similar cases that several peptide sequences expressed on bacteriophages and selected for the functionality do not function when they are prepared as peptides themselves, nor the sequences are impossible to express as soluble peptides. An expression system as a peptide-fusion with Escherichia coli maltose binding protein (MBP) was established. This system has been reported effective in improving the above mentioned problem. The MBP gene was cloned for this purpose, and an expression vector was newly constructed. The 18mer linear peptide and 11mer cyclic peptide was successfully expressed on MBP and 18mer but not 11mer peptide reacted with the mAb used. Less
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