Molecular interactions: oral bacteria & matrix proteins

革兰氏阴性菌与牙周病有关，牙周病是一组牙龈和牙周组织支持结构的慢性炎症性疾病。放线杆菌伴生放线杆菌是一种革兰氏阴性兼性球菌，主要分布于口腔和上呼吸道。这种细菌与年轻人的牙周炎和成人牙周炎密切相关。这种病原体与其他严重的人类感染有关，如心内膜炎、软组织脓肿，以及最近的心血管疾病。虽然牙周组织被认为是这些口外感染的来源，但人们对AA在口腔内维持自身以及在组织中渗透和传播所使用的取向知之甚少。病原体已经进化出不同的策略，成功地在宿主组织上定植。这些病原体的一个共同主题是，在严酷或恶劣的条件下，通过与特定宿主大分子的黏附来启动感染的能力。这些分子包括宿主细胞分泌的形成细胞外基质的蛋白质。这种基质通常由胶原和特定的非胶原蛋白组成，例如纤维连接蛋白。在牙周组织中发现的主要蛋白质是胶原，我们已经证明AA既能与胶原结合又能与纤维连接蛋白结合。在这项建议中，我们计划确定编码基质结合蛋白的基因，并确定这些蛋白结合所需的氨基酸序列。我们的假设是，基质结合蛋白的合成参与了牙周袋和底层组织的AA定植。为了实现这些目标，我们建议1)通过构建转座子突变和噬菌体展示文库来分离胶原和纤维连接蛋白结合蛋白的编码基因；2)确定基因序列并产生等基因突变体；3)确定与结合活性有关的蛋白区域(S)；4)确定LJP患者血清对这些蛋白的免疫反应性。这一信息可用于牙周病疫苗治疗剂的开发。

Gram-negative bacteria are associated with periodontal diseases, which are a group of chronic inflammatory diseases of the gingiva and the supporting structures of the periodontium. Actinobacillus actinomycetemcomitans (Aa) is a Gram-negative, facultative coccobacillus that colonizes the human oral cavity and upper respiratory tract. This bacterium is closely associated with periodontitis in young individuals and with cases of adult periodontitis. This pathogen has been associated with other serious human infections such as endocarditis, soft tissue abscesses, and more recently cardiovascular disease. Although the periodontium is believed to be the source of these extraoral infections, little is known about the tropisms used by Aa to maintain itself within the oral cavity and to infiltrate and disseminate in tissues. Pathogens have evolved diverse strategies to be successful in colonization of the host tissue. A common theme amongst these pathogens is the ability to initiate infection by adhesion to specific host macromolecules under stringent or hostile conditions. These molecules include proteins that are secreted by host cells that form the extracellular matrix. This matrix is usually composed of collagen and specific noncollagenous proteins, e.g. fibronectin. The major protein found in the periodontium is collagen and we have demonstrated that Aa binds to both collagen and fibronectin. In this proposal, we plan to identify the genes coding for matrix binding proteins and determine the amino acid sequences of these proteins required for binding. It is our hypothesis that the synthesis of matrix binding proteins is involved in Aa colonization of the periodontal pocket and underlying tissues. In order to realize these goals, we propose to 1) isolate the genes coding for collagen and fibronectin binding proteins by constructing a transposon mutagenesis and a phage display library; 2) determine the gene sequences and generate isogenic mutants; 3) determine the regions(s) of the protein essential for binding activity; 4) determine the immunoreactivity of LJP patient sera to these proteins. This information can be used for the development of therapeutic agents of vaccines for periodontal disease.