Secretin Architecture

分泌素架构

• 批准号: 9245656
• 负责人: MICHAEL S DONNENBERG
• 金额: $ 19.19万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9245656
• 关键词: Architecture; Bacteria; Biochemical; Biogenesis; Biological Models; Biomedical Research; Cells; Complex; Cryoelectron Microscopy; Cysteine; Development; Disease; Disulfides; Electron Microscopy; Energy Transfer; Equipment; Exploratory/Developmental Grant; Fiber; Fimbriae Proteins; Goals; Homologous Gene; Human; Imagery; Ion Channel; Knowledge; Length; Maps; Membrane; Microscopic; Modeling; Mutagenesis; Pilum; Positioning Attribute; Proteins; Publishing; Research; Research Project Grants; Resolution; Secretin; Shapes; Site; Structure; Surface; System; Techniques; Technology; Toxin; Virulence Factors; X-Ray Crystallography; crosslink; disulfide bond; electron tomography; enteropathogenic Escherichia coli; experimental study; fluorophore; improved; in vivo; monomer; mutant; novel; novel therapeutics; periplasm; permissiveness; protein complex; public health relevance; tool

 DESCRIPTION (provided by applicant): The secretin, a homomultimeric outer membrane channel, is the only protein common to type IV pilus (T4P), type 2 secretion and type 3 secretion systems. Therefore, secretins enjoy a unique position of prominence among virulence factors. Although the structures of several periplasmic secretin domains have been solved by X-ray crystallography, and the overall shape of secretins has been described by electron microscopy, sufficient detail to understand the mechanism that allows secretins to interact with other components of their respective machines and to open selectively to allow passage of their substrates has not be achieved. In this proposal we describe how we will advance our understanding of secretin function. Using a variety of techniques, including cysteine mutagenesis and Förster resonance energy transfer, we described a topology model for BfpB, the secretin from the T4P of enteropathogenic Escherichia coli. The first aim of this proposal will build on that knowledge by using in vivo disulfide cross-linking to trap the pilin in the secretin and thereby map the walls of the secretin channel. Recent advances in cryo-electron microscopy have permitted the structural analysis of large complexes at resolutions approaching the atomic range. In aim two we will capitalize on these advances to visualize the secretin and proteins with which it associates in unprecedented detail. The experiments described in this proposal will provide critical structural information that is essential for our lng-term goal of a complete understanding of T4P biogenesis. Furthermore, this information will also have implications for secretion systems and may ultimately have practical implications for new therapeutics.

 描述（由适用提供）：Systrin，一种同源外膜外膜通道，是IV型Pyrus（T4P），类型2分泌和3型分泌系统的唯一蛋白质。因此，Secitins在病毒因素中享有独特的突出地位。尽管已经通过X射线晶体学求解了几个周围秘密蛋白结构域的结构，并且通过电子显微镜描述了Secitins的整体形状，足以了解允许秘密与其各自机器的其他组件相互作用的机制，并可以选择性地进行允许允许其底物的通过。在此提案中，我们描述了如何提高我们对Secralin功能的理解。使用多种技术，包括半胱氨酸诱变和Förster共振能量转移，我们描述了BFPB的拓扑模型，该模型是肠病毒性大肠杆菌T4P的Systin。该提议的第一个目标将 通过使用体内二硫键交联来将PILIN捕获到Sectialin中，从而绘制Sectionin通道的墙壁，从而建立在这些知识的基础上。冷冻电子显微镜的最新进展允许在接近原子范围的分辨率下对大型复合物进行结构分析。在目标两个方面，我们将利用这些进步，以形象化其与其前所未有的细节相关的分泌素和蛋白质。本提案中描述的实验将提供关键的结构信息，这对于我们对T4P生物发生的完全了解的LNG期目标至关重要。此外，这些信息还将对分泌系统产生影响，并最终可能对新的治疗学产生实际影响。