RECONSTITUTION OF PILUS BIOGENESIS IN A CELL-FREE SYSTEM

侵袭病原体的第一步是附着。 给它的主人。依恋通常是通过毛发状的附属物来调节的 从细菌表面辐射出来，称为菌毛。了解皮卢斯 组装将提供一条重要的防御细菌的途径- 相关的疾病，并将提供对基本的 细胞器生物发生的生物学过程。这项建议旨在 利用Defined在无细胞系统中重建菌毛生物发生 组件。泌尿系致病性大肠埃希菌P菌毛的研究 模型皮卢斯，将用于重建。P菌毛很复杂 至少由六个不同的子单元组成的结构，并且需要两个 特化组装蛋白：外膜引导器和周质 监护人。外膜引导器将被结合到脂质体中， 然后菌毛亚单位会被困在里面或添加到外面 这些蛋白脂质体。伴侣-亚单位复合体与细胞的结合 Usher蛋白、菌毛亚结构的组装和完整的组装 将对菌毛进行检查。成功重建菌毛生物发生将 为分析突变体和确定 亚基和组装蛋白的功能。重组将会 促进对菌毛-宿主细胞相互作用的理解，并将 协助设计和测试抗菌剂。

The first step in the invasion of a pathogenic bacterium is attachment to its host. Attachment is often mediated by hairlike appendages that radiate out from the bacterial surface, called pili. Understanding pilus assembly will provide an important avenue of defense against bacterial- related diseases, and will also provide insight into the fundamental biological process of organelle biogenesis. This proposal aims to reconstitute pilus biogenesis in a cell-free system using defined components. P pili from uropathogenic Escherichia coli, a well studied model pilus, will be used for reconstitution. P pili are complex structures composed of at least six different subunits, and require two specialized assembly proteins: an outer membrane usher and a periplasmic chaperone. The outer membrane usher will be incorporated into liposomes, and pilus subunits will then be trapped inside or added to the outside of these proteoliposomes. Binding of chaperone-subunit complexes to the usher protein, assembly of pilus substructures, and assembly of complete pili will be examined. Successful reconstitution of pilus biogenesis will provide a powerful tool for analyzing mutants and determining the function of the subunits and assembly proteins. Reconstitution will facilitate the understanding of pilus-host cell interactions, and will aid in the design and testing of antimicrobial agents.