Characterization of Heat Shock Proteins from Periodontopathogenic Bacteria and Their Virulences

牙周病原菌热激蛋白的表征及其毒力

• 批准号: 08672372
• 负责人: HINODE Daisuke
• 金额: $ 1.47万
• 依托单位: The University of Tokushima
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08672372/
• 关键词: Heat shock protein; Periodontopathogenic bacteria; Immunological characterization; Purification; Specific antibody; Bacteroides forsythus; Porphromonas gingivalis; Actinobacillus actinomycetemcomitans; 熱ショックタンパク質; 歯周病原細菌; 歯周病原性

Recently, considerable attention has been given to the potential roles of heat shock protein (HSP) of in inflammation and autoimmune diseases. In this study, we established the isolation procedure of HSP from periodontopathogenic bacteria and investigated the immunological characterization of them using the specific antibodies.The isolation procedure of HSPs from whole cells of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Bacteroides forsythus was performed by affinity chromatography on adenosine 5'-triphosphate-agarose followed by preparative polyacrylmide gel electrophoresis and could obtain the considerable amount of purified GroEL- and DnaK-like proteins. Also, these proteins could be successfully used to obtain rabbit sera which possessed high titer of antibodies and a very good specificities comparing to the commercial ones.By Western-immunoblotting, it found that the GroEL-like protein from P.gingivalis was localized in cytoplasm and the DnaK-like protein f … More rom P.gingivalis was localized in both cytoplasm and periplasm.Analysis of the N-terminal amino acid sequence of each DnaK-like protein from P.gingivalis and B.forsythus showed a high degree of homology with the DnaK protein from Escherichia coli.However, these two HSPs reacted very weakly with a commercial anti-DnaK polyclonal antibody by dot-blotting.GroEL-like proteins isolated P.gingivalis, A.actinomycetemcomitans and B.forsythus showed a high degree of homology of their N-terminal amino acid sequences. Polyclonal antibodies raised against each GroEL-like protein showed a high level of cross-reactivity. Reactivity of these antibodies against recombinant human HSP60 was weak.Our findings suggest that DnaK- and GroEL-like proteins from periodontopathogens are well conserved and that the GroEL-like protein resemble each other more closely. It is possible to suggest that HSPs from bacteria are important targets of the immune response, however, these anti-infectious immune response may shift from their own protection to pathogenesis by their molecular mimicry. Less

近年来，热休克蛋白(HSP)在炎症和自身免疫性疾病中的潜在作用受到了广泛关注。本研究建立了牙周病致病菌热休克蛋白的分离方法，并用特异性抗体对其进行了免疫学性质的研究。通过5‘-三磷酸-腺苷-琼脂糖亲和层析及制备性聚丙烯酰胺凝胶电泳法从牙龈卟啉单胞菌、伴生放线杆菌和福赛斯类杆菌的全细胞中分离热休克蛋白，可获得大量纯化的GroEL-和DNAK-like蛋白。免疫印迹结果表明，牙龈假单胞菌的GroEL样蛋白定位于细胞质中，DNAK样蛋白f…定位于细胞质中牙龈假单胞菌和连翘假单胞菌的DNAK蛋白的N端氨基酸序列分析表明，它们与大肠杆菌的DNAK蛋白有很高的同源性，但这两个热休克蛋白与商品化的抗DNAK多克隆抗体反应很弱。针对每种GroEL样蛋白的多克隆抗体表现出高度的交叉反应。这些抗体对重组人HSP60的反应性较弱。我们的发现表明来自牙周病原体的DNAK和GroEL样蛋白被很好地保存，并且GroEL样蛋白彼此更接近。细菌热休克蛋白可能是免疫应答的重要靶点，然而，这些抗感染免疫应答可能通过其分子模拟而从自身保护转变为致病。较少

项目成果

Hinode, Daisuke et al.: "A general procedure for the isolation of heat shock proteins from periodontopathogenic bacteria" J.Microbiol. Methods. 25. 349-355 (1996)

Hinode、Daisuke 等人：“从牙周病原细菌中分离热休克蛋白的一般程序”J.Microbiol。

Hinode, Daisuke et al.: "Cross-reactivity of Specific antibodies directed to that shock proteins from periodontopathogenic bacteria and of human origin" Oral Microbiol.Immunol.13. 55-58 (1998)

Hinode、Daisuke 等人：“针对牙周病原细菌和人类来源的休克蛋白的特异性抗体的交叉反应性”Oral Microbiol.Immunol.13。

日野出大輔、他5名: "Porphyromonas gingivalis の熱ショック蛋白質" 口腔衛生学会雑誌. 47. 412-413 (1997)

Daisuke Hinode 等 5 人：“牙龈卟啉单胞菌的热休克蛋白”，口腔健康学会杂志 47. 412-413 (1997)。

Hinode,D.,Grenier,D.and Mayrand,D.: "A general procedure for the isolation of heat-shock proteins from periodontopathogenic bacteria" Journal of Microbiological Methods. 25. 349-355 (1996)

Hinode,D.、Grenier,D. 和 Mayrand,D.：“从牙周病原细菌中分离热休克蛋白的一般程序”微生物方法杂志。

Hinode, D.et al.: "Heat shock proteins from Porphyromonas gingivalis" J.Dental Health. 47. 412-413 (1997)

Hinode, D.等人：“牙龈卟啉单胞菌的热休克蛋白”J.Dental Health。