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（由申请人提供）：III型分泌（T3 S）系统是一种多组分转运装置，介导细菌鞭毛和多种病原体使用的毒性相关注射体的组装。分泌是由质子动力驱动的。FlhE是鞭毛T3 S系统的成员，其缺失导致E中的质子泄漏。大肠杆菌和沙门氏菌，伴随细胞死亡。在所有的T3 S系统中，有一组六个保守的整合膜（IM）蛋白分泌所必需的。尽管存在这种保守性，但尚不清楚这些蛋白质中哪些实际上构成分泌孔，哪些仅起支持作用。该提议试图利用FlhE在限定分泌孔中的性质。在急诊大肠杆菌/沙门氏菌鞭毛系统，FlhA-FlhB被推测构成了“输出门”。FlhE作为flhBAE操纵子的一部分转录，因此可能在FlhAB功能中起作用。FlhE是周质蛋白，包括在鞭毛基体内腔内。我们从周质中分离出FlhE并解析了其结构。1.5A的分辨率结构表明FlhE可能作为分泌孔的塞子。以FlhE结构为指导，我们建议测试FlhE如何充当插头。这些实验将确定分泌通道及其支撑支架的成分。相似的整体架构 鞭毛和injectisomes的基本结构，其几个T3 S组分蛋白的保护，以及在其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是可转移到其他。由于缺乏FlhE会导致大量的细胞死亡，拟议的研究可能会提出合理的药物设计策略，针对肠道病原体，FlhE主要是found. In长期，我们预计，从这些研究中获得的知识可以用于靶向PMF驱动的多药外排泵或其他必要的质子驱动的分泌通道。