FlhE as a probe for the flagellar Type III secretion pore

FlhE 作为鞭毛 III 型分泌孔的探针

• 批准号: 8698613
• 负责人: Rasika M Harshey
• 金额: $ 23.18万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-15 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8698613
• 关键词: Architecture; Caliber; Cell Death; Complex; Drug Design; Drug Targeting; Enterobacteriaceae; Escherichia coli; Flagella; Genetic; Housing; Integral Membrane Protein; Investigation; Knowledge; Link; Mediating; Membrane; Molecular Chaperones; Mutagenesis; Operon; Pathogenesis; Periplasmic Proteins; Physiological; Play; Property; Protein Secretion; Proteins; Proton-Motive Force; Protons; Resolution; Role; Salmonella; Structure; Substrate Specificity; Surface; System; Testing; Therapeutic Intervention; Virulence; cell motility; efflux pump; enteric pathogen; kinetosome; member; pathogen; periplasm; public health relevance; research study; scaffold; secretion process

DESCRIPTION (provided by applicant): The Type III secretion (T3S) system is a multicomponent transport apparatus that mediates the assembly of both the bacterial flagellum and the virulence-associated injectisomes used by a wide variety of pathogens. Secretion is driven by proton motive force. FlhE is a member of the flagellar T3S system, whose absence causes a proton leak in E. coli and Salmonella, concomitant with cell death. In all T3S systems, there is a set of six conserved integral membrane (IM) proteins essential for secretion. Despite this conservation, it is not known which of these proteins actually constitute the secretion pore, and which play only supportive roles. This proposal seeks to exploit the properties of FlhE in defining the secretion pore. In the E. coli/Salmonella flagellar system, FlhA-FlhB are speculated to comprise the 'export gate'. FlhE is transcribed as part of the flhBAE operon, hence likely plays a role in FlhAB function. FlhE is a periplasmic protein, included within the flagellar basal body lumen. We isolated FlhE from the periplasm and solved its structure. The 1.5 A resolution structure suggests that FlhE may serve as a plug for the secretion pore. With the FlhE structure as guide, we propose to test how FlhE acts as a plug. These experiments will identify the constituents of the secretion channel and its supporting scaffold. The similar overall architecture of the basal structure of flagella and injectisomes, conservation of several of their T3S component proteins, as well as similarities in their PMF-driven secretion mechanism, suggests that lessons obtained from one system are transferable to the other. Because absence of FlhE causes substantial cell death, the proposed studies could potentially suggest rational drug-design strategies for targeting enteric pathogens where FlhE is mainly found. In the long-term, we anticipate that the knowledge gained from these studies could be used to target PMF-driven multidrug efflux pumps or other essential proton-driven secretion channels.

描述(申请人提供)：III型分泌物(T3S)系统是一种多组分的运输装置，它调节多种病原体使用的细菌鞭毛和与毒力相关的注射体的组装。分泌是由质子动力驱动的。FlhE是鞭毛状T3S系统的一员，它的缺失会导致大肠杆菌和沙门氏菌中的质子泄漏，并伴随细胞死亡。在所有的T3S系统中，都有一组对分泌至关重要的保守的整合膜(IM)蛋白。尽管有这种保守性，但目前尚不清楚这些蛋白质中的哪些实际上构成了分泌孔，哪些只起到了辅助作用。这项提议试图利用FlhE的特性来定义分泌孔。在大肠杆菌/沙门氏菌鞭毛系统中，FlhA-FlhB被推测构成“出口门”。FlhE被转录为flhBAE操纵子的一部分，因此可能在FlhAB功能中发挥作用。FlhE是一种周质蛋白，包括在鞭毛基体腔内。我们从周质中分离出FlhE并解决了它的结构问题。1.5A的分辨率结构表明，FlhE可能是分泌孔的塞子。以FlhE结构为指导，我们建议测试FlhE作为插头的作用。这些实验将确定分泌通道及其支撑支架的成分。类似的整体架构 鞭毛和注射体的基本结构、它们几个T3S组成蛋白的保守以及它们由PMF驱动的分泌机制的相似性表明，从一个系统中获得的经验教训可以移植到另一个系统中。由于缺乏FlhE会导致大量细胞死亡，拟议的研究可能会建议针对主要发现FlhE的肠道病原体的合理药物设计策略。从长远来看，我们预计从这些研究中获得的知识可以被用于靶向PMF驱动的多药外排泵或其他基本的质子驱动的分泌通道。