EPITOPE MAPPING OF BLACK-PIGMENTED BACTEROIDES FIMBRIAE

黑色拟杆菌菌毛的表位作图

• 批准号: 3875388
• 负责人: ROBERT J GENCO
• 金额: --
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3875388
• 关键词: Actinomyces; Bacteroides gingivalis; bacterial antigens; bacterial pigment; cell adhesion; conformation; crosslink; epitope mapping; genetic library; germ free condition; human subject; immunochemistry; laboratory rat; membrane activity; membrane proteins; microorganism culture; microorganism immunology; monoclonal antibody; periodontitis; pilus; protein structure; proteolipids; receptor; western blottings

The long-term objective of this study is to explore which fimbrial epitopes of black-pigmented Bacteroides mediate colonization to eukaryotic cell surfaces. Specifically, the fimbrial proteins from oral black-pigmented Bacteroides, including B. gingivalis, B. intermedius, and B. intermedius, and B. endodontalis, will be purified, and a library of monoclonal antibodies against their natural epitopes will be prepared. These monoclonal reagents will be tested for their ability to inhibit fimbrial attachment to epithelial cells. This will permit us to identify fimbrial epitopes of biological relevance. These epitopes will then be characterized with regard to their primary peptide structure and conformation. Next, the peptides of important regions of the fimbrial molecules will be synthesized and their immunogenicity with various carriers will be determined in an attempt to elicit antibodies which would inhibit fimbrial mediated epithelial attachment. The synthetic peptides will then be coupled to iodinatable heterobifunctional crosslinking agents to identify receptors on epithelial cells which mediate fimbrial attachment. Biochemical and immunochemical methods will be used with an emphasis on production and analysis of synthetic peptides. The functional assays include evaluation of the binding of black-pigmented Bacteroides to epithelial cells. The significance of these findings resides in the potential importance of fimbriae in the colonization of black-pigmented Bacteroides. Fimbriae are likely responsible for colonization by interacting with epithelial cells, other bacteria in coaggregates, and may be important in interacting with the host protective cells, such as neutrophils. Once these virulence-mediating functions have been elucidated, and the domains of the fimbriae responsible for these interactions identified, design of vaccines using synthetic peptides of critical domains could be considered. Thus, the studies outlined in this subproject could then provide the groundwork for a rational approach for immunization to modulate colonization of black-pigmented Bacteroides.

本研究的长期目标是探索哪种菌毛 表位的黑色色素类杆菌介导定植， 真核细胞表面。 具体来说，来自口腔黑色素沉着的纤毛蛋白 拟杆菌属，包括B。gingivalis，B. B. intermedius和B. intermedius和B.牙髓，将被纯化，和一个文库， 针对其天然表位的单克隆抗体将被 制备 将检测这些单克隆试剂的 抑制菌毛附着于上皮细胞的能力。 这将 允许我们鉴定生物相关的菌毛表位。 然后，这些表位将关于它们的表位特征进行表征。 一级肽结构和构象。 接下来， 菌毛分子的重要区域将被合成 并确定其与各种载体的免疫原性 为了引发抗体， 介导的上皮附着。 然后，合成肽将 与可碘化的异双官能交联剂偶联， 鉴定上皮细胞上介导菌毛的受体 附件. 生物化学和免疫化学方法将 用于生产和分析合成 缩氨酸 功能测定包括结合的评价， 黑色素拟杆菌的上皮细胞。 这些发现的意义在于， 菌毛在黑色素沉着菌定植中的重要性 拟杆菌 菌毛可能是负责殖民 与上皮细胞，其他细菌在共聚集体， 并且在与宿主保护性细胞相互作用中可能是重要的 细胞，如中性粒细胞。 一旦这些调解服从的 功能已经阐明，和领域的菌毛 负责这些相互作用确定，疫苗的设计， 可以考虑使用关键结构域的合成肽。 因此，该分项目中概述的研究可以提供 为合理的免疫接种方法奠定基础， 黑色拟杆菌的定植。