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Molecular biological analysis of virulent factors on P. gingivalis related with periodontal disease

Research Project

Project/Area Number 08457485
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Morphological basic dentistry
Research InstitutionKanagawa Dental College,

Principal Investigator

UMEMOTO Toshio  Kanagawa Dental College, Oral Microbiology, Professor, 歯学部, 教授 (20067036)

Co-Investigator(Kenkyū-buntansha) TAKAHASHI Yusuke  Kanagawa Dental College, Oral Microbiology, Assistant, 歯学部, 助手 (20267511)
HAMADA Nobushiro  Kanagawa Dental College, Oral Microbiology, Assistant, 歯学部, 助手 (20247315)
YOSHIMOTO Hisashi  Kanagawa Dental College, Oral Microbiology, Assistant Professor, 歯学部, 助教授 (60084787)
Project Period (FY) 1996 – 1998
Project Status Completed (Fiscal Year 1998)
Budget Amount *help
¥6,900,000 (Direct Cost: ¥6,900,000)
Fiscal Year 1998: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1997: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 1996: ¥3,000,000 (Direct Cost: ¥3,000,000)
KeywordsPeriodontopathic bacteria / Porphyromonas gingivalis / Fimbriae / firmA / Transformation / Adhirence factor / Surface hydrophobicity / Coaggregation / 線毛 / 形質転換 / 細胞付着性 / 線毛遺伝子 / fimA不活化株
Research Abstract

Genetic approaches have been recently introduced to analyze virulence factors of Porphyromonas gingivalis.
In this study, a recombinant plasmid pYHF1, containing a gene firmA 381 encoding the main subunit of the fimbriae of P. gingivalis 381 was constructed by ligating a linear pUC13Bg12.1 fragment into pYH420, and was successfully electroporated into YH522, a restriction-deficient P. gingivalis host strain constructed in our previous study. Several wild, restriction-positive P. gingivalis strains, including O-131, W50, BLO-1, BH18/10 and ATCC 33277, also accepted pYHF1 when the plasmid DNA purified from P. gingivalis YH522 was used as the donor, albeit at an extremely low frequency. Among these host strains, O-131, W50 and BLO-1, exhibited a difference in the fimbrial antigenicity from strain 381 as well as YH522, although the other 2 strains, BH18/10 and ATCC 33277, exhibited the same antigenicity as 381.
Production of a specific protein with a molecular weight of 41-kDa was observed b … More y SDS-PAGE in all fimA transformants irrespective of the host strain. This product was considered to be the recombinant fimbrillin, because it reacted with a polyclonal antiserum raised against the fimbriae of ATCC 33277, a type strain of P. gingivalis containing the same fimbrial antigenicity as 381. In addition, all transformants except strain ATCC 33277 exhibited characteristic fimbrial structures (recombinant fimbriae), which were distinguishable by electron microscopy from their native fimbriae, although a marked difference was observed in the amount of gene expression as shown by the density of the protein bands. Moreover, an apparent relationship between the amounts of the recombinant fimbrillin and the recombinant fimbriae was also observed.
When then compared various biological properties such as cell-surface hydrophobicity, ability of attachment to the epithelial cells, co-aggregation with other bacteria, and hemagglutination activity between the transformants containing the recombinant fimbriae and the host cells without fimbriae. In all cases where increased expression of the recombinant fimbriae was clearly observed, the transformants exhibited reduced attachment ability, coaggregation and hydrophobicity. No difference in hemagglutination activitiy, however, was observed between any combination of the firmA-containing and non-containing cells.
These results suggested that the recombinant fimbriae produced by the firmA gene have some unknown difference in their biological natures from the native fimbriae, possibly due to the lack of some minor components which are essential for co-aggregation with other bacteria and attachment to epithelial cells. Less

Report

(4 results)
  • 1998 Annual Research Report   Final Research Report Summary
  • 1997 Annual Research Report
  • 1996 Annual Research Report
  • Research Products

    (5 results)

All Other

All Publications (5 results)

  • [Publications] Yoshimoto H.,Takahashi Y.,Kato D.,and Umemoto T: "Construction of a plasmid vector for transformation of Porphyromonas gingivalis."FEMS Microbiology Letters,. 152. 175-181 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] 鈴木正之,浜田信城,高橋祐介: "Porphyromonas gingivalis.染色体DNAの制限酵素地図および遺伝子地図の作製"神奈川歯学. 32. 216-226 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Takahashi Y,Kato D,Hamada N,Yoshimoto H,Umemoto T: "Transformation and Expression of a cloned FimA Gene in Porphyromonas gingivalis."Infection and Immunity. 67. 2013-2018 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Hisashi Yoshimoto, Yusuke Takahashi, Daisuke Kato, and Toshio Umemoto: "Construction of a plasmid vector for transformation of Porphyromonas gingivalis"FEMS Microbiology Letters. 152. 175-181 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Yusuke Takahashi, Daisuke Kato, Nobushiro Hamada, Hisashi Yoshimoto, Toshio Umemoto: "Transformation and Expression of a cloned Fim A Gene in Porphyromonas gingivalis"Infect Immun. 67. I2013-2018 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary

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Published: 1996-04-01   Modified: 2016-04-21  

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