Fibronectin-binding Protein of Treponema Denticola in Periodontal Disease

牙周病中齿螺旋体的纤连蛋白结合蛋白

• 批准号: 8191607
• 负责人: XIAOPING XU
• 金额: $ 22.28万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8191607
• 关键词: Adherence; Adhesions; Adhesives; Adult; Affect; Anaerobic Bacteria; Antibodies; Bacteria; Bacterial Adhesins; Bacterial Adhesion; Bacterial Infections; Bacteroides forsythus; Binding; Binding Proteins; Binding Sites; Biological; Biological Process; Cell surface; Cell-Matrix Junction; Cells; Characteristics; Clostridium difficile; Databases; Disease; Escherichia coli; Extracellular Matrix; Family; Fibrinogen; Fibronectins; Gene Family; Gene Proteins; Genes; Genetic; Genome; Glycoproteins; Goals; Human; Infection; Knowledge; Laminin; Liquid substance; Mediating; Molecular; Mus; N-terminal; Order Spirochaetales; Pathogenesis; Pathogenicity; Peptides; Periodontal Diseases; Periodontal Infection; Periodontitis; Peritoneal; Phenotype; Plasma; Play; Porphyromonas gingivalis; Property; Protein Family; Proteins; Random Peptide Libraries; Recombinants; Research; Role; Screening procedure; Site-Directed Mutagenesis; Streptococcus pneumoniae; Streptococcus pyogenes; Structural Protein; Structure-Activity Relationship; Testing; Tissues; Treponema denticola; Virulence; Virulence Factors; Western Blotting; base; design; in vivo Model; innovation; knockout gene; member; microorganism; mortality; mouse model; mutant; novel; pathogen; prevent; research study; surface coating; synthetic peptide

DESCRIPTION (provided by applicant): Periodontitis is the second most common disease worldwide and is induced by microorganisms including Treponema denticola. Fibronectin (FN) is a multifunctional adhesive glycoprotein that is distributed in plasma, the extracellular matrix, and on the surface of cells. Bacterial FN-binding proteins play important roles in bacterial adhesion and invasion of host tissues. Previous research has found that T. denticola bound FN which was blocked by anti-FN antibodies. However, the knowledge about FN-binding proteins and their roles in the pathogenesis of T. denticola is very limited. The genome project of T. denticola has identified a new putative fibronectin/fibrinogen-binding protein gene, fbp (Gene ID: 2739776), which is homologous to an N-terminal segment of the prokaryotic FN-binding protein. (FbpA, pfam05833). FbpA is a family of homologous Fn-binding protein gene with over 117 members from different bacteria according to the CDD database. The role of fbp in pathogenesis of T. denticola and the interactions between Fbp and FN has not been investigated. In preliminary studies, we expressed recombinant Fbp in E. coli, and identified that rFbp has FN binding properties. The binding of T. denticola to FN was blocked by rFbp in a concentration dependent manner to a maximum level of 42%. We also make anti-rFbp antibodies to confirm that Fbp is expressed in T. denticola by Western blot. We propose that fbp is an important virulence factor. The research hypothesis is that Fbp contributes to the virulence of T. denticola by mediating interaction with host FN. The purpose of this proposal is to analyze the virulence roles of fbp by comparing the phenotypes of fbp mutant and wildtype strains, and to characterize the functions of Fbp by studying the interactions between Fbp protein and FN. Specifically, in Aim 1 we plan to make isogenic mutants of fbp in T. denticola. We will compare fbp mutant with wildtype T. denticola in FN binding, cell attachment, and virulence in a mouse model. In addition, we will rescue fbp mutants by genetic complementation to confirm the function of fbp. In Aim 2, we will screen a random peptide library to identify the binding site of Fbp for FN and subsequently verify the binding site by site-specific mutagenesis on fbp. In addition we will also design and test the capacity of synthetic Fbp peptide to inhibit the binding of T. denticola to FN. We expect to find a peptide which prevents the adherence of T. denticola to host tissue which may serve as an anti-infection factor. In summary, this is first adhesin mutant of T. denticola and first systemic approach to identity the binding site of Fbp protein. We believe that the proposed project is novel and unique and will help us to understand not only the role of fbp in the pathogenesis of T. denticola, but also in other bacteria with fbp genes. The Long term goal is to understand in depth the mechanism of attachment and invasion of T. denticola to periodontal tissues and to characterize the biological function of the FbpA protein family in order to provide new strategies to prevent bacterial infections. PUBLIC HEALTH RELEVANCE: Periodontitis is the second most common disease worldwide and is associated with infection by the bacteria Treponema denticola. This study will investigate pathogenicity of a recently identified fibronectin binding protein gene (fbp) of T. denticola as a basis for new strategies to prevent bacterial infection and periodontal disease.

描述（由申请人提供）：牙周炎是全球第二常见的疾病，是由包括treponema denticola在内的微生物引起的。纤连蛋白（FN）是一种多功能粘合剂糖蛋白，分布在血浆，细胞外基质和细胞表面上。细菌FN结合蛋白在细菌粘附和宿主组织侵袭中起重要作用。先前的研究发现，被抗FN抗体阻断的牙齿牙霉结合的FN。然而，关于FN结合蛋白及其在牙霉菌发病机理中的作用的知识非常有限。 T. denticola的基因组项目已经鉴定出一种新的推定纤连蛋白/纤维蛋白原结合蛋白基因FBP（基因ID：2739776），该基因与促促性FN结合蛋白的N末端段同源。 （FBPA，PFAM05833）。根据CDD数据库，FBPA是一个同源FN结合蛋白基因的家族，来自不同细菌的117个成员。尚未研究FBP在牙霉菌发病机理中的作用以及FBP和FN之间的相互作用。在初步研究中，我们表达了大肠杆菌中的重组FBP，并确定RFBP具有FN结合特性。牙霉菌与FN的结合以浓度依赖性方式的RFBP阻断，最大水平为42％。我们还制作抗RFBP抗体，以确认FBP通过Western blot在T. denticola中表达。我们建议FBP是重要的毒力因素。 研究假设是，FBP通过介导与宿主FN的相互作用来促进牙霉菌的毒力。该建议的目的是通过比较FBP突变体和野生型菌株的表型来分析FBP的毒力作用，并通过研究FBP蛋白与FN之间的相互作用来表征FBP的功能。具体而言，在AIM 1中，我们计划使T. denticola中的FBP的等生突变体。我们将在Fn结合，细胞附着和小鼠模型中的FN牙杆菌与牙齿的野生型突变体进行比较。此外，我们将通过遗传互补来挽救FBP突变体，以确认FBP的功能。在AIM 2中，我们将筛选一个随机肽库，以识别FBP的FBP结合位点，并随后通过位点特异性诱变在FBP上验证结合位点。此外，我们还将设计和测试合成FBP肽的能力，以抑制牙霉菌与FN的结合。我们期望找到一种肽，该肽可以防止牙霉菌对托管组织的粘附，这可能是抗感染因子。 总而言之，这是牙霉菌的第一个粘附素突变体，也是FBP蛋白的结合位点的第一种全身方法。我们认为，拟议的项目是新颖且独特的，它将不仅有助于我们了解FBP在T. denticola的发病机理中的作用，而且还可以理解其他具有FBP基因的细菌。长期目标是深入了解牙霉菌对牙周组织的依恋和侵袭的机理，并表征FBPA蛋白家族的生物学功能，以提供防止细菌感染的新策略。 公共卫生相关性：牙周炎是全球第二常见的疾病，与细菌Treponema denticola感染有关。这项研究将研究牙霉菌的最近确定的纤连蛋白结合蛋白基因（FBP）的致病性，作为预防细菌感染和牙周疾病的新策略的基础。