Interactions of the oral pathogen, A. actinomycetemcomitans, with collagen

描述(申请人提供)：牙周病是一组炎症性疾病，导致牙龈和牙齿支撑结构的破坏。组织的丢失是由于特定的牙龈下细菌的定植引起的宿主免疫反应的失衡造成的。这种微生物区系也被认为是口腔外疾病的来源。伴生放线杆菌是一种与慢性和局部侵袭性牙周疾病相关的革兰氏阴性牙周病原体。多种口腔外播散性疾病可归因于这种病原体，包括感染性心内膜炎和心血管疾病的加重。然而，伴生放线菌定植于口腔或渗入并扩散到远处组织的嗜性仍然难以捉摸。这些组织共享共同的细胞外基质(ECM)蛋白，包括胶原。我们的团队已经发现了一种新的蛋白质，细胞外基质蛋白粘附素A(EmaA)，它对细菌与胶原蛋白的相互作用至关重要，也是感染性心内膜炎启动的毒力决定因素。总之，伴生放线杆菌表达一种细胞表面粘附素，该粘附素与胶原结合，胶原是最丰富的细胞外基质蛋白，用于建立感染源。因此，我们推测，细胞外基质的定植是牙周沟再次感染和扩散到外周的蓄水池。 口腔网站。EmaA粘附素由三个相同的单体组成，这些单体是放线菌伴生菌独有的，它们嵌入外膜，组装独特的天线状表面附件，对胶原结合至关重要。然而，这些单体的蛋白质大小和序列是血清型特有的。此外，EmaA被O-多糖(O-PS)糖翻译后修饰，O-PS糖是与胶原结合的关键。有趣的是，这种EmaA修饰与O-PS生物合成所需的酶相同。糖链在与胶原蛋白相互作用中的具体作用尚不清楚。此应用程序的目标是 目的：阐明该病原菌在口腔定植并向其他组织扩散的机制(S)。为了实现这一目标，我们提出了以下具体目标：1)从b型细菌中确定EmaA的糖链部分的定位和组成；2)确定EmaA基因和细菌血清型在胶原结合中的作用；以及3)EmaA/胶原蛋白相互作用的生化和生物物理测定。确定EmaA活性结合部分的性质将有助于理解该蛋白在伴生放线杆菌发病机制中的作用。从这项研究中获得的对这种粘附素分子机制的新见解将导致开发分子来破坏和/或减少这种病原体的组织定植。

DESCRIPTION (provided by applicant): Periodontal disease represents a group of inflammatory diseases, which lead to the destruction of the gingiva and the supporting structures of the teeth. The loss of tissue results from an imbalance of the host immune response induced by the colonization of specific sub-gingival bacteria. This microflora is also considered the source for extra-oral diseases. Aggregatibacter actinomycetemcomitans is a Gram-negative periodontal pathogen associated with both the chronic and localized aggressive forms of the disease. Multiple disseminated extra-oral diseases are attributed to this pathogen including infective endocarditis and the potentiation of cardiovascular disease. However, the tropism used by A. actinomycetemcomitans to colonize the oral cavity or infiltrate and disseminate to distant tissues has remained elusive. These tissues share common extracellular matrix (ECM) proteins including collagen. Our group has identified a novel protein, extracellular matrix protein adhesin A (EmaA), which is critical for the interaction of the bacterium with collagens and is a proven virulence determinant for the initiation of infective endocarditis. In summary, A. actinomycetemcomitans expresses a cell surface adhesin that binds collagen, the most abundant ECM protein, for establishing infectious foci. Therefore, we posit that ECM colonization acts as a reservoir for re-infection of the gingival sulcus and dissemination to extra oral sites. The EmaA adhesin is composed of three identical monomers unique to A. actinomycetemcomitans that intercalate in the outer membrane to assemble unique antenna-like surface appendages essential for collagen binding. However, the protein size and sequence of the monomers is serotype specific. Furthermore, EmaA is posttranslationally modified with O-polysaccharide (O-PS) sugars that are crucial for binding to collagen. Interestingly, this EmaA modification shares the same enzymes required for O-PS biosynthesis. The specific role of the glycan moieties in the interaction with collagen remains undefined. The goal of this application is to elucidate the mechanism(s) utilized by this pathogen for colonizing the oral cavity and disseminating to other tissues. To achieve this goal we propose the following specific aims: 1) determine the localization and composition of the glycan moiety of EmaA from serotype b bacteria; 2) determine the role of EmaA genotype and bacterial serotype in collagen binding; and 3) biochemical and biophysical determination of the EmaA/collagen interaction. Defining the nature of the active binding moiety of EmaA will aid in understanding the role of this protein in the pathogenesis of A. actinomycetemcomitans. The new insights gained from this study into the molecular mechanism of this adhesin will lead to the development of molecules to disrupt and/or diminish tissue colonization by this pathogen.