Protein Interactions in the Flagellar Export Machinery

描述（由申请人提供）：运动性在细菌发病中起关键作用。许多物种利用鞭毛推进来定位寄主和在组织中移动。鞭毛是一种复杂的、可逆的、自组装的纳米机器。它有自己的III型蛋白质输出机制，与毒力因子分泌机制密切相关。出口装置的膜蛋白可能是鞭毛中最不为人所知的成分。拟议研究的总体目标是获得鼠伤寒沙门氏菌这些蛋白质之间相互作用的详细知识。此外，了解鞭毛组装的最终目标之一是重新组装鞭毛输出复合体，以确定和分析组成蛋白的作用以及机械本身。为了实现提案的具体目标并接近长期目标，将进行几项努力：通过亲和印迹、共凝固等生化方法，鉴定出口装置内膜蛋白（FIhA、FlhB、FliO、FliP、FliQ和FliR）之间的相互作用以及膜组分与其他鞭毛蛋白之间的相互作用；生物物理分析与分析性超离心和荧光共振能量转移研究的相互作用；出口器械蛋白复合物的重构及其协同功能分析。

DESCRIPTION (provided by applicant): Motility plays a critical role in bacterial pathogenesis. Many species utilize flagella propulsion to locate hosts and move through tissues. The flagellum is a complex, reversible, self-assembling nanomachine. It contains its own type III protein export machinery that is closely related to the virulence factor secretion apparatus. The membrane proteins of the export apparatus are probably the least understood components of the flagellum. The overall objective of the proposed research is to obtain detailed knowledge of the interactions among these proteins from Salmonella typhimurium. Additionally, one of the ultimate goals of efforts to understand flagellar assembly is to reassemble the flagellar export complex from components to determine and analyze the roles of constituent proteins as well as the machinery itself. In order to achieve the specific objectives of the proposal and to approach the long term goal, several efforts will be undertaken: identification of interactions among the membrane proteins within the export apparatus (FIhA, FlhB, FliO, FliP, FliQ and FliR) and between the membrane components and other flagellar proteins by biochemical methods such as affinity blotting, copurification, etc.; biophysical analysis of interactions with analytical ultracentrifugation and fluorescence resonance energy transfer studies; and reconstitution of complexes of export apparatus proteins and analysis of their concerted function.