BINDING OF KLEBSIELLA FIMBRIAE TO RESPIRATORY TISSUE

肺炎克雷伯菌的3型纤维粘附素（MrkD）介导细菌与特定类型胶原蛋白的结合。这种粘附素分子的受体位于组织的基底膜区域，脱落的上皮表面可能存在结合位点。由于肺炎克雷伯菌是上皮损伤患者呼吸道感染的常见原因，由于MrkD粘附素的存在，3型毛状菌可能在这些个体的气道中定植。在本提案中，我们描述了实验，我们描述了实验，以确定MrkD粘附素在促进呼吸道体内定植中的作用。将使用呼吸道感染的小鼠模型，并用稀释的HCI或六甲基磷酰胺治疗动物，以引起粘膜脱屑，以模仿在受损个体中看到的类似组织学病变。一个MrkD阳性的肺炎克雷伯菌分离株，以及该菌株不表达MrkD粘附素的衍生物，将用于研究粘附素在定植中的作用。为了确定这些分子内的结合结构域，将进行变异MrkD粘附蛋白的分子分析。将构建具有两种具有受体结合特异性的不同分子的特定区域的嵌合粘附素。结合到特定的胶原类型将通过标准技术进行研究。位点定向诱变将用于进一步描述粘连素的胶原结合位点。此外，胶原亚基和片段将被检查，以便在整个胶原蛋白分子上定位受体位点。该材料将用于几种结合试验，使用纯化的毛状蛋白和毛状细菌。受体位点的鉴定将允许进一步研究开发可以抑制敏感个体细菌定植的合成类似物。

The type 3 fimbrial adhesin (MrkD) of Klebsiella pneumoniae mediates binding of the bacteria to specific types of collagen. The receptors for this adhesin molecule are located in the basement membrane region of tissues, and denuded epithelial surfaces are likely to present binding sites. Since K. pneumoniae is a frequent cause of respiratory infections of patients with epithelial trauma, it is possible that type 3 fimbriate bacteria colonize the airways in these individuals due to the presence of the MrkD adhesin. In this proposal we describe experiments we describe experiments to determine the role of the MrkD adhesin in facilitating colonization, in vivo, of the respiratory tract. A murine model of respiratory infection will be used and the animals will be treated with dilute HCI or hexamethylphosphoramide to cause desquamation of the mucosa to mimic similar histological lesions seen in compromised individuals. An mrkD-positive isolate of K. pneumoniae, and derivatives of this strain that do not express the MrkD adhesin, will be used to investigate the role of the adhesin in colonization. Molecular analysis of variant MrkD adhesin proteins will be performed in order to determine the binding domains within these molecules. Chimeric adhesins possessing defined regions of two different molecules exhibiting receptor binding specificities will be constructed. Binding to specific collagen types will be investigated by standard techniques. Site-directed mutagenesis will be used to further delineate the collagen-binding sites of the adhesin. Also, collagen subunits and fragments will be examined in order to localize the receptor sites on whole collagen molecules. This material will be used in several binding assays using purified fimbrial protein as well as fimbriate bacteria. The identification of receptor sites will allow further investigations into the development of synthetic analogs that could inhibit bacterial colonization of susceptible individuals.