Surface protein complexes of oral treponemes: assembly and host cell interactions

口腔密螺旋体的表面蛋白复合物：组装和宿主细胞相互作用

• 批准号: 7464047
• 负责人: J CHRISTOPHER FENNO
• 金额: $ 36.58万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-05 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7464047
• 关键词: Affect; Anabolism; Animal Model; Bacteria; Bacterial Proteins; Behavior; Binding; Biochemical; Cell Communication; Cell Surface Receptors; Cells; Cellular Assay; Chronic; Complex; Environment; Enzymes; Epithelial Cells; Epithelial Receptor Cell; Event; Extracellular Matrix; Genetic; Gingiva; Goals; Human; Individual; Infection; Inflammatory; Knowledge; Laboratories; Lead; Lesion; Lipoproteins; Mediating; Mediator of activation protein; Membrane; Membrane Proteins; Microbe; Molecular Analysis; Molecular Biology; Mutation; Nutrient; Operon; Oral; Order Spirochaetales; Organism; Penetration; Peptide Hydrolases; Periodontal Diseases; Permeability; Positioning Attribute; Process; Protein Binding; Proteins; Proteolytic Processing; Public Health; Qualifying; Refractory; Reporting; Research; Role; Series; Surface; TLR2 gene; Testing; Tissues; Treponema denticola; acquired immunity; cytotoxic; dentilisin; genetic analysis; insight; interest; member; oral spirochetes; polypeptide; protein complex; receptor; response

DESCRIPTION (provided by applicant): The goal of this research is to characterize expression and assembly of Treponema denticola (Td) surface proteins that mediate interactions with host tissue, thereby gaining insight into mechanisms of periodontal disease. We focus on analysis of two Td protein complexes that directly affect host cells: the oligomeric Msp protein and the PrtP lipoprotein protease complex. The overall hypothesis is that Msp and PrtP are major contributors to Td cytopathic behavior in periodontal disease. Expression of Msp and the PrtP complex are interdependent. To understand their roles in microbe-host interactions, we must understand how these molecules are expressed and interact in the spirochete. We propose genetic analysis of structural features and interactions of these complexes and their components. Our approach is to define the mechanisms of interdependence between Msp and the PrtP complex in order to characterize their roles in microbe-host interactions. Aim 1: Determine requirements for assembly and activity of the PrtP complex. We will construct a series of specific mutations in the protease locus that will enable us to characterize posttranslational processing of PrcA and PrtP, expression of PrcB, localization of protease complex proteins and assembly of the active protease complex. Aim 2: Determine the role of Msp in biosynthesis of protease complex proteins, and the role of the protease in oligomerization of Msp. We will characterize interactions between Msp and protease operon proteins that influence expression and activity of each outer membrane complex. Aim 3: Characterize interactions of Td outer membrane components with epithelial cells. To further studies of host cell responses to Td challenge, we will characterize interactions between Msp and a putative receptor identified on epithelial cells and characterize early events in epithelial cell innate responses to the PrtP protease complex and Msp, with the goal of differentiating between roles of Msp and the protease complex in cellular responses to Td. This project, involving genetic and biochemical analyses of Td outer membrane complex expression and analysis of Td effects on host cells, will contribute to molecular analysis of microbe-host interactions in the oral environment. Our laboratory is uniquely positioned to conduct these studies. Completion of these Aims will contribute to both basic knowledge of spirochete molecular biology and to understanding of microbe-host interactions in chronic infections such as periodontal diseases. Public Health Significance: This project will provide information on how a key bacterium involved in initiation periodontal disease interacts with human cells. The studies will first characterize the process of protein complex assembly on the bacterial surface. Then we will determine which specific proteins on human cells these complexes bind to, and then assay cellular responses to bacterial protein complexes binding to cell surface receptors.

描述（由申请人提供）：本研究的目标是表征介导与宿主组织相互作用的齿垢密螺旋体 (Td) 表面蛋白的表达和组装，从而深入了解牙周病的机制。我们重点分析两种直接影响宿主细胞的 Td 蛋白复合物：寡聚 Msp 蛋白和 PrtP 脂蛋白蛋白酶复合物。总体假设是 Msp 和 PrtP 是牙周病 Td 细胞病变行为的主要贡献者。 Msp 和 PrtP 复合物的表达是相互依赖的。为了了解它们在微生物与宿主相互作用中的作用，我们必须了解这些分子如何在螺旋体中表达和相互作用。我们建议对这些复合物及其成分的结构特征和相互作用进行遗传分析。我们的方法是定义 Msp 和 PrtP 复合物之间的相互依赖机制，以表征它们在微生物-宿主相互作用中的作用。目标 1：确定 PrtP 复合物的组装和活性的要求。我们将在蛋白酶基因座中构建一系列特定突变，这将使我们能够表征 PrcA 和 PrtP 的翻译后加工、PrcB 的表达、蛋白酶复合物蛋白的定位以及活性蛋白酶复合物的组装。目标 2：确定 Msp 在蛋白酶复合物蛋白生物合成中的作用，以及蛋白酶在 Msp 寡聚化中的作用。我们将表征 Msp 和蛋白酶操纵子蛋白之间的相互作用，这些相互作用影响每个外膜复合物的表达和活性。目标 3：表征 Td 外膜成分与上皮细胞的相互作用。为了进一步研究宿主细胞对 Td 攻击的反应，我们将表征 Msp 和上皮细胞上鉴定的假定受体之间的相互作用，并表征上皮细胞对 PrtP 蛋白酶复合物和 Msp 固有反应的早期事件，目的是区分 Msp 和蛋白酶复合物在细胞对 Td 反应中的作用。该项目涉及 Td 外膜复合物表达的遗传和生化分析以及 Td 对宿主细胞的影响分析，将有助于口腔环境中微生物与宿主相互作用的分子分析。我们的实验室具有独特的优势来开展这些研究。完成这些目标将有助于掌握螺旋体分子生物学的基础知识，以及了解牙周病等慢性感染中微生物与宿主相互作用的知识。公共卫生意义：该项目将提供有关引发牙周病的关键细菌如何与人体细胞相互作用的信息。这些研究将首先描述细菌表面蛋白质复合物组装的过程。然后我们将确定这些复合物与人类细胞上的哪些特定蛋白质结合，然后分析细胞对与细胞表面受体结合的细菌蛋白质复合物的反应。