Molecular and biological studies of colonization factors of oral spirochetes

口腔螺旋体定植因子的分子生物学研究

• 批准号: 11671830
• 负责人: UMEMOTO Toshihiko
• 金额: $ 2.3万
• 依托单位: Asahi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671830/
• 关键词: oral spirochetes; colonization factors; coaggregation; chemotaxis; albumin-binding proteins; flagellar genes; 細菌定着因子; 表層タンパク質; Treponema medium; Porphyromonas gingivalis; 分離精製

The aim of this poject was to investigate molecular and biological analyses of colonization factors of human oral spirochetes. The Porphyromonas gingivalis fimbria-binding polypeptide, fragellar genes, and albumin-binding polypeptides of a medium-sized Treponema medium ATCC 700293 were focused.1. Immunoelectron microscopy demonstrated that the 37 kDa fimbria-binding polypeptide was localized on the outer envelope. The surface protein was partially purified from SDS solubilized cells by the use of several chromatography matrices, including affinity chromatography on a polyclonal anti-37 kDa polypeptide serum-CNBr-activated Sepahrose 4B matrix. The purified protein had an IpH of 7.4, and exhibited the property of binding to the fimbria from P.gingivalis ATCC 33277 and 381.2. The DNA fragment of the T.medium was cloned and sequenced. The size of the fragment was 15,809 bp including flagellar genes flgD and flgE.The flgD sequence was 480 bp in length, and flgE sequence (873 bp) was separat … More ed with a transposon-like sequence.. The nucleotide sequence of T.medium flgD and flgE genes were relatively preserved when compared with other host-associated treponemal species.3. The motility and chemotaxis of human oral spirochetes Treponema denticola ATCC 35404, T.medium, and T.vincentii ATCC 35580 were examined by a capillary assay method. Of five tested sera three human oral treponemes were chemoattractant to rabbit serum. The checkerboard analysis of chemotaxis toward the rabbti serum showed that T.denticola cells swam toward the culutre media containing higher concentrations of the serum. The treponemal chemotaxis was reduced by heating serum, and increased by addition of rabbit albumin. Western blotting analysis demonstrated that these treponemes possessed rabbit albumin-binding polypeptides with molecular sizes of 65 kDa and 70 kDa. Immunoelectron microscopy demonstrated that the 65 kDa rabbit albumin-binding polypeptide was located on their outer envelopes, suggesting that the rabbit albumin-binding polypeptide seem to be responsible for chemotaxis toward rabbit serum. Less

本课题旨在从分子水平和生物学水平研究口腔螺旋体定植因子。研究了中等大小密螺旋体ATCC 700293培养基的牙龈卟啉单胞菌菌毛结合多肽、碎片基因和白蛋白结合多肽。免疫电子显微镜显示，37 kDa的菌毛结合多肽位于外被膜上。通过使用几种色谱基质，包括在多克隆抗37 kDa多肽血清-CNBr-活化Sepahrose 4 B基质上的亲和色谱，从SDS溶解的细胞中部分纯化表面蛋白。纯化的蛋白质具有7.4的IpH，并表现出与来自牙龈卟啉单胞菌ATCC 33277和381.2的菌毛结合的性质。克隆了培养基中的DNA片段并进行了测序。该片段大小为15，809 bp，包含鞭毛基因flgD和flgE，其中flgD序列长480 bp，flgE序列长873 bp。 ...更多信息 艾德与转座子一样的序列。与其他宿主相关密螺旋体相比，该菌株flgD和flgE基因的核苷酸序列相对保守.用毛细管电泳法检测了人口腔螺旋体齿垢密螺旋体ATCC 35404、培养基密螺旋体和文森特密螺旋体ATCC 35580的运动性和趋化性。在5份供试血清中，3份人口腔密螺旋体对兔血清具有化学引诱作用。棋盘格法分析表明，齿垢锥虫细胞向含较高浓度血清的培养基中游动。加热血清可降低密螺旋体的趋化性，而加入兔白蛋白则可增强其趋化性。Western blotting分析表明，这些密螺旋体具有兔白蛋白结合多肽的分子大小为65 kDa和70 kDa。免疫电子显微镜显示，65 kDa的兔白蛋白结合多肽位于它们的外包膜，这表明兔白蛋白结合多肽似乎是负责对兔血清的趋化性。少

项目成果

T.Umemoto: "Fimbria-mediated coaggregation between human oral anaerobes Treponema medium and Porphyromonas gingivalis."Microbiology and Immunology. 43(9). 837-845 (1999)

T.Umemoto：“人口腔厌氧菌密螺旋体培养基和牙龈卟啉单胞菌之间菌毛介导的共聚集。”微生物学和免疫学。

T.Umemoto: "Finbria-mediated coaggregation between human oral anaerobes Treponema medium and Porphyromoans gingivalis"Microbiologyand Immunology. 43(9). 837-845 (1999)

T.Umemoto：“Finbria 介导的人口腔厌氧菌密螺旋体介质和牙龈卟啉菌之间的共聚集”微生物学和免疫学。

Toshihiko Umemoto, Fuminobu Yoshimura, Hidetoshi Kureshiro, Jun-ichiro Hayashi, Toshihide Noguchi, and Tomohiko Ogawa: "Fimbria-mediated coaggregation between human oral anaerobes Treponema medium and Porphyromonas gingivalis."Microbiology and Immunology.

Toshihiko Umemoto、Fuminobu Yoshimura、Hidetoshi Kureshiro、Jun-ichiro Hayashi、Toshihide Noguchi 和 Tomohiko Okawa：“人口腔厌氧菌密螺旋体介质和牙龈卟啉单胞菌之间菌毛介导的共聚集。”微生物学和免疫学。