EPITOPE MAPPING OF BLACK-PIGMENTED BACTEROIDES FIMBRIAE

这项研究的长期目标是探索哪种菌毛 黑色素类杆菌抗原表位介导的定植 真核细胞表面。 具体地说，口腔黑色素菌毛蛋白 拟杆菌，包括牙龈假单胞菌、中间假单胞菌和假单胞菌。 Intermedius和B.endodontalis将被提纯，并建立一个 针对其天然表位的单抗将是 准备好了。这些克隆试剂将进行测试，以确定它们的 抑制菌毛附着于上皮细胞的能力。这将是 允许我们确定具有生物相关性的菌毛表位。 然后，这些表位将根据它们的 初级多肽的结构和构象。下一步，多肽 菌毛分子的重要区域将被合成 并将测定它们与各种载体的免疫原性。 试图诱导出抑制菌毛的抗体 中介上皮附着。然后合成的多肽将 偶联到可碘的异双官能团交联剂上 确定上皮细胞上介导菌毛的受体 依恋。生化和免疫化学方法将是 主要用于合成纤维的生产和分析。 多肽。功能分析包括结合的评估 从黑色素类杆菌到上皮细胞。 这些发现的意义在于潜在的 菌毛在黑色素细菌定植中的重要性 拟杆菌属。菌毛很可能是由 与上皮细胞、聚集体中的其他细菌相互作用， 并且在与宿主保护性的相互作用中可能很重要 细胞，如中性粒细胞。一旦这些毒力调停 功能已被阐明，菌毛的结构域 负责这些相互作用的确定，疫苗的设计 可以考虑使用临界区的合成肽。 因此，这一分项目概述的研究可以提供 为免疫调节的合理方法奠定基础 黑色素类杆菌的定植。

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.