CRYO-EM TOMOGRAPHY OF FLAGELLAR FILAMENTS OF BORRELIA BURGDORFERI

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 中心，不一定是研究者的机构。 摘要： 螺旋体是一种医学上重要的细菌，但对它的了解却很少。 这些微生物引起多种重要疾病，包括梅毒、莱姆病、回归热和钩端螺旋体病。它们也与牙周病和人类牙周病有关。 研究人员的目的是回答有关螺旋体周质鞭毛（PFs）的结构和运动的某些问题。 螺旋体的鞭毛是所有细菌鞭毛中最复杂的。 在PF的表面上是由一到两种FlaA蛋白种类组成的蛋白质鞘。 核心由三到四种蛋白质的聚合物组成，称为FlaB蛋白。单个PF蛋白的功能尚不清楚。 在以前的工作与RVBC，分离的PFs突变体鞭毛进行了研究，冷冻EM，手稿正在准备中。 在这项研究中，我们希望通过比较具有改变的运动性的各种突变株来证实来自完整细胞的鞭毛中的发现。 这些研究特别相关，因为鞭毛和/或运动性被认为是螺旋体和其他几种致病菌的毒力因子。 在上一个报告期内，将四种伯氏疏螺旋体菌株作为冷冻断层扫描的完整标本进行了急冻：B31野生型、Che A、Che X和Fli G1。 对所有四种菌株进行断层扫描重建。 Charon博士访问了资源研究结果，并在电子显微镜下观察标本，以帮助确定未来工作的最佳范例。 我们发现原生细胞中鞭毛的排列与以前在化学固定细胞中看到的不同，并且与以前研究的随机薄切片相比，在整个细胞的断层图像中可以更好地理解运动装置的3-D排列。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. ABSTRACT: Spirochetes are a medically significant but a remarkable poorly understood group of bacteria. These organisms cause a variety of important diseases including syphilis, Lyme disease, relapsing fever, and leptospirosis. They are also implicated in periodontal disease and human diarrheal disease. The investigators aim to answer certain questions relating to the structure of spirochete periplasmic flagella (PFs) and motility. Spirochete PFs are the most complex of any bacterial flagellar filament. On the surface of the PFs is a protein sheath comprised of one to two FlaA protein species. The core is comprised of a polymer of three to four protein species referred to as FlaB proteins. The function of the individual PF proteins is not understood. In previous work with the RVBC, isolated PFs from mutant flagella were studied by cryo-EM, and a manuscript is in preparation. In this study, we wish to confirm findings in flagella from intact cells, by comparing various mutant strains having altered motility. These studies are especially relevant in that flagella and/or motility are implicated as a virulence factor for spirochetes and several other species of pathogenic bacteria. In the previous reporting period, four strains of Borrelia burgdorferi were plunge-frozen as whole mounts for cryo-tomography: B31 wild type, Che A, Che X, and Fli G1. Tomographic reconstructions of all four strains were made. Dr. Charon visited the Resource to study the results, and to observe the specimens at the electron microscope in order to help identify the optimal examples for future work. We found the arrangement flagella in native cells to differ from that seen previously in chemically-fixed cells, and the 3-D arrangement of the motile apparatus could be much better appreciated in the tomograms of whole cells, compared to the random thin sections previously studied.