Structural mechanisms of Clostridium difficile pathogenesis

艰难梭菌发病机制的结构机制

基本信息

批准号：
9212765
负责人：
Dana Borden Lacy
金额：
$ 39.43万
依托单位：
VANDERBILT UNIVERSITY MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-05-15 至 2021-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9212765
关键词：
Anaerobic Bacteria Antibiotics Antibodies Apoptosis Bacteria Binding Binding Sites CRISPR/Cas technology Cell physiology Cells Cellular Assay Clostridium difficile Colon Complex Crystallization Cytosol Data Development Diarrhea Disease Electron Microscopy Endosomes Epithelial Cells Event Family Fluorescence Funding Genetic Variation Glucosyltransferase Goals Guanosine Triphosphate Phosphohydrolases Homologous Protein Hospitals Human Human poliovirus Hybrids Hydrophobicity Incidence Individual Infection Inflammation Integration Host Factors Intoxication Label Lead Measurement Mediating Membrane Modeling Molecular Mus Mutagenesis Mutation NADPH Oxidase Necrosis Normal tissue morphology Pathogenesis Pathology Physiological Prevention Production Proteins Pseudomembranous Colitis Public Health Publishing Reactive Oxygen Species Reproduction spores Research Role Series Severities Structure Testing Therapeutic Therapeutic Intervention Tissues Toxic Megacolon Toxin United States Vaccine Antigen Vaccines Variant Viral Proteins Virulence Factors X-Ray Crystallography alpha Toxin alpha helix cost crosslink cytotoxicity design electron tomography experimental study follow-up human disease improved mouse model new therapeutic target novel therapeutics poliovirus receptor programs public health relevance receptor receptor mediated endocytosis rho scaffold therapeutic target tool

项目摘要

DESCRIPTION (provided by applicant): Clostridium difficile is a gram-positive, spore-forming anaerobe that infects the colon, causing a range of human disease including diarrhea, pseudomembranous colitis, and toxic megacolon. The incidence, severity, and costs associated with C. difficile infection (CDI) are increasing, making C. difficile a significant public health concern. The principle virulence factors in C. difficile pathogenesis are TcdA and TcdB, two large homologous toxins capable of entering and modifying multiple targets within eukaryotic host cells. This proposal is designed to follow up two important discoveries made in the previous funding cycle, a crystal structure of TcdA and the cellular receptor for TcdB. In Aim 1, we will use the structure of the TcdA delivery domain as a framework for understanding toxin pore formation within the host endosomal membrane. A hybrid approach of electron microscopy, crosslinking, and fluorescence intensity measurements will generate a structural view of the toxin pore. In Aim 2, we will define the interaction of TcdB with its receptor, PVRL3, using a hybrid of electron microscopy, mutagenesis, quantitative binding studies, and X-ray crystallography. In Aim 3, we will evaluate the relevance of the TcdB-PVRL3 interaction in mouse models of intoxication and infection. Results from these aims will provide a molecular understanding of the events that allow toxins access to the host cell and a framework for therapeutic intervention.

描述（由申请人提供）：艰难梭菌是一种革兰氏阳性、产芽孢厌氧菌，可感染结肠，引起一系列人类疾病，包括腹泻、伪膜性结肠炎和中毒性巨结肠。与艰难梭菌相关的发病率、严重程度和费用。艰难梭菌感染 (CDI) 正在增加，使艰难梭菌成为重要的公共卫生问题。艰难梭菌发病机制的主要毒力因素是。 TcdA 和 TcdB 是两种大型同源毒素，能够进入并修饰真核宿主细胞内的多个靶标。该提案旨在跟进上一个资助周期中的两项重要发现，即 TcdA 的晶体结构和 TcdB In 的细胞受体。目标 1，我们将使用 TcdA 传递结构域的结构作为了解宿主内体膜内毒素孔形成的框架，一种电子显微镜的混合方法。交联和荧光强度测量将生成毒素孔的结构视图。在目标 2 中，我们将结合使用电子显微镜、诱变、定量结合研究和 X 射线来定义 TcdB 与其受体 PVRL3 的相互作用。在目标 3 中，我们将评估 TcdB-PVRL3 相互作用在小鼠中毒和感染模型中的相关性，这些目标的结果将提供对事件的分子理解。毒素进入宿主细胞和治疗干预的框架。