Mechanism of the Usher in Assembly and Secretion of Pili

描述(申请人提供)：病原菌必须组装并 分泌毒力因子，以便与宿主相互作用并引起 疾病。革兰氏阴性细菌除了有一个外膜外，还有一个外膜。 细胞质膜，必须通过它们分泌毒力因子 障碍。发生这种情况的机制可能非常复杂，并不是 很好理解。伴侣/引导者途径是一种毒力蛋白分泌。 需要两种成分才能通过外膜分泌的途径 周质伴侣和称为引座者的外膜蛋白。这个 伴侣引导分泌蛋白质的正确折叠，并防止其 参与非生产性互动。引座员是一个集合 位于外膜的平台，为细胞提供分泌通道 浮出水面。伴侣/引座者途径是组装和分泌 广泛存在于革兰氏阴性菌中的粘附性结构超家族 病原体。由该途径组装的原型细胞器是P和 尿路致病性大肠埃希菌表达的1型菌毛 尿路感染的病原体。P菌毛和1型菌毛是临界毒力 因素，允许肾和膀胱的结合和定植， 分别进行了分析。该提案的长期目标是通过以下方式使用菌毛生物发生 致尿性大肠杆菌作为了解毒力的模式系统 革兰氏阴性杆菌的因子分泌。更具体地说，结构 并将对引座员的功能进行研究，以阐明分子 控制外膜分泌的机制。第一个具体 目的是创建一个详细的引座员结构安排模型 外膜使用计算机分析和表位测绘技术。这个 第二个具体目的是通过生成和生成来探讨引座员的功能 对突变体的分析。第三个具体目标是建立一个无牢房系统 对于菌毛的生物发生，基于将引导者重新构建为脂质体。是这样的 一个系统将为研究监护人/引座员提供一个宝贵的工具 途径和分析突变体。本提案中描述的工作将 阐明毒力因子分泌机制并创造机会 不仅治疗尿路的新型抗菌剂的开发 感染，但也有广泛的传染病。

DESCRIPTION (provided by the applicant): Pathogenic bacteria must assemble and secrete virulence factors in order to interact with their hosts and cause disease. Gram-negative bacteria have an outer membrane in addition to a cytoplasmic membrane and must secrete virulence factors across both these barriers. The mechanisms by which this occurs can be quite complex and are not well understood. The chaperone/usher pathway is a virulence protein secretion pathway that requires two components for secretion across the outer membrane: a periplasmic chaperone and an outer membrane protein termed an usher. The chaperone directs proper folding of the secreted proteins and prevents their engagement in non-productive interactions. The usher serves as an assembly platform at the outer membrane and provides a secretion channel to the cell surface. The chaperone/usher pathway is required for assembly and secretion of a superfamily of adhesive structures in a broad range of Gram-negative pathogens. The prototypical organelles assembled by this pathway are the P and type 1 pili expressed by uropathogenic Escherichia coli, the primary causative agent of urinary tract infections. P and type 1 pili are critical virulence factors, allowing binding and colonization of the kidney and bladder, respectively. The long-term goal of this proposal is to use pilus biogenesis by uropathogenic E. coli as a model system with which to understand virulence factor secretion in Gram-negative bacteria. More specifically, the structure and function of the usher will be investigated to elucidate the molecular mechanisms governing secretion across the outer membrane. The first specific aim is to create a detailed model of the structural arrangement of the usher in the outer membrane using computer analysis and epitope mapping techniques. The second specific aim is to probe function of the usher through generation and analysis of mutants. The third specific aim is to establish a cell-free system for pilus biogenesis based on reconstitution of the usher into liposomes. Such a system will provide an invaluable tool for studying the chaperone/usher pathway and analyzing mutants. The work described in this proposal will elucidate mechanisms of virulence factor secretion and create opportunities for the development of novel antimicrobial agents to treat not only urinary tract infections, but also a broad range of infectious diseases.