Genetic Analysis-Adhesion of S Sanguis to S Pellicle

描述：先前的拨款支持导致发现了一种新的副血链球菌菌毛相关的粘附，Fapl。参与Fapl糖基化的Fapl样分子和基因在整个口腔链球菌中是惊人地保守的，并且也在重要的病原体如肺炎链球菌和金黄色葡萄球菌中发现。我们将使用S。parasanguis作为模型系统来研究这些病原菌中Fap 1样蛋白的功能。Fapl是S.副血吸虫生物膜我们已经确定Fapl（rFapl）的N-末端多肽对于细菌初始附着到生物膜表面是必需的。Fapl是糖基化的，并且该蛋白质的糖基化参与生物膜的成熟。本申请的总体目标通过两个具体目的突出：（1）确定初始细菌附着所需的rFapl粘附素的结构/功能关系和（2）定义Fapl糖基化和细菌生物膜形成中fapl侧翼基因的功能。我们将把遗传学，结构生物学（NMR光谱学和X射线晶体学），体外结合生化测定和体内动物模型研究结合在一起，以解决Fapl多肽的结构如何影响其功能。这些全面的方法应该阐明Fapl与唾液受体结合的结构基础，以及预测其他医学上重要的病原体的Fapl样分子。fapl侧翼区的蛋白质可能构成Fapl的偶联分泌和糖基化途径。为了开始理解所涉及的机制，我们将确定该位点中蛋白质的亚细胞定位，并鉴定体内和体外假定的蛋白质-蛋白质相互作用。我们先前证明Fapl糖基化参与生物膜的成熟。使用分子和遗传分析相结合，我们将剖析这些糖基化基因在生物膜形成中的功能。Fapl糖基化功能的阐明和Fapl多肽结构/功能关系的定义将有助于口腔环境中细菌定植的领域和其他重要病原体的致病机制，因为它们的基因组表明它们也具有Fapl样分子和侧翼蛋白。

DESCRIPTION: Previous grant support has led to the discovery of a novel Streptococcus parasanguis fimbriae associated adhesion, Fapl. Fapl-like molecules and genes involved in Fapl glycosylation are strikingly conserved throughout oral streptococci and are also found in important pathogens, such as Streptococcus pneumoniae and Staphylococcus aureus. We will use S. parasanguis as a model system to study the function of Fapl-like proteins in these pathogenic bacteria. Fapl is required for the formation of a S. parasanguis biofilm. We have determined that the N-terminal polypeptide of Fapl (rFapl) is essential for initial attachment of bacteria to the biofilm surface. Fapl is glycosylated, and glycosylation of this protein is involved in maturation of biofilm. The overall goal of this application is highlighted by two Specific Aims: (1) to determine the structure/function relationship of the rFapl adhesin that is required for initial bacterial attachment and (2) to define the functions of the fapl flanking genes in Fapl glycosylation and bacterial biofilm formation. We will bring genetic, structural biology (NMR spectroscopy and X-ray crystallography), in vitro binding biochemical assays and in vivo animal model studies together to address how the structure of the Fapl polypeptide influences its function. These comprehensive approaches should shed light on the structural basis for binding of Fapl to salivary receptors, and for predicted Fapl-like molecules of other medically important pathogens. Proteins of the fapl flanking region may constitute a coupled secretion and glycosylation pathway for Fapl. To begin to understand the mechanisms involved, we will determine the subcellular Iocalization of proteins in this locus and identify putative protein-protein interactions in vivo and in vitro. We previously demonstrated that Fapl glycosylation is involved in maturation of the biofilm. Using a combination of molecular and genetic analyses, we will dissect the function of these glycosylation genes in biofilm formation. Elucidation of the function of Fapl glycosylation and definition of the Fapl polypeptide structure/function relationships will contribute to the field of bacterial colonization in the oral environment and to the pathogenic mechanism of other important pathogens as their genomes indicate that they also possess Fapl-like molecules, and the flanking proteins.