Structure-Function Relationships in the Spirochetal Flagellar Motor

摘要 螺旋体是一组在遗传学上不同的细菌，对人类健康具有重要意义 因为它们会引起重大疾病，如梅毒（梅毒螺旋体），莱姆病（伯氏疏螺旋体）， 钩端螺旋体病（问号钩端螺旋体）和牙周炎（密螺旋体属）。感染并传播 作为哺乳动物宿主，螺旋体已经进化出一种独特的形态和运动性， 通过粘性介质和组织屏障转移。螺旋体运动所必需的细胞器 是周质鞭毛，其存在于细菌周质空间中并且与外部鞭毛不同。 模型系统中的鞭毛大肠杆菌和肠道沙门氏菌。考虑到鞭毛驱动的运动是 对于致病性螺旋体和许多其他细菌的毒力至关重要，我们的长期目标是了解 鞭毛组装和功能的分子机制。在上一个财政年度，我们 证明莱姆病螺旋体B. burgdorferi（Bb）是一个很好的模型系统， 周质鞭毛原位在一个前所未有的决议。与Md Motaleb博士和Chunhao博士合作 Li，我们已经产生并表征了包括60种不同鞭毛和趋化性的大型Bb文库 变种人在了解周质鞭毛及其显著的 驱动独特的螺旋体运动和形态的能力。本申请的目的是阐明 周质鞭毛的三个基本但具有挑战性的方面：1）鞭毛的结构和功能 Ⅲ型分泌器; 2）质子动力驱动鞭毛旋转的机制 鞭毛通过改变旋转方向来控制运动的机制 和趋化性。与遗传和生物化学方法一起，冷冻ET将用于确定 纳米尺度下自然细胞环境中螺旋体鞭毛马达的结构/功能关系 分辨率

ABSTRACT Spirochetes are a phylogenetically distinct group of bacteria that are of significant importance in human health as they cause major diseases such as syphilis (Treponema pallidum), Lyme disease (Borrelia burgdorferi), leptospirosis (Leptospira interrogans), and periodontitis (Treponema spp.). To infect and disseminate in mammalian hosts, spirochetes have evolved a unique morphology and motility that is highly effective at translocating through viscous media and tissue barriers. The organelles essential for spirochetal motility are periplasmic flagella, which reside in the bacterial periplasmic space and are distinct from the external flagella in the model systems Escherichia coli and Salmonella enterica. Given that flagella-driven motility is crucial for virulence of pathogenic spirochetes and many other bacteria, our long-term goal is to understand molecular mechanisms underlying flagellar assembly and function. During the previous funding period, we have demonstrated that the Lyme disease spirochete B. burgdorferi (Bb) is a great model system for characterizing periplasmic flagella in situ at an unprecedented resolution. In collaboration with Drs. Md Motaleb and Chunhao Li, we have generated and characterized a large Bb library including 60 different flagellar and chemotaxis mutants. Significant progress has been made in understanding the periplasmic flagella and their remarkable capacity in driving the unique spirochetal motility and morphology. The objective of this application is to illuminate three fundamental but challenging aspects of the periplasmic flagella: 1) the structure and function of the flagellar type III secretion apparatus; 2) the mechanism underlying the flagellar rotation driven by proton motive force across membrane; and 3) the mechanisms by which flagella switch rotational directions to control the motility and chemotaxis. Together with genetic and biochemical approaches, cryo-ET will be utilized to determine the structure/function relationship of the spirochetal flagellar motor in a native cellular environment at nanometer resolution.