STREPTOCOCCAL PLATELET BINDING AND ENDOCARDITIS

链球菌血小板结合和心内膜炎

基本信息

批准号：
2673029
负责人：
PAUL M. SULLAM
金额：
$ 23.47万
依托单位：
NORTHERN CALIFORNIA INSTITUTE/RES/EDU
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-07-01 至 2001-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2673029
关键词：
Streptococcus sanguis bacterial endocarditis bacterial genetics bacterial proteins cell adhesion cell adhesion molecules human tissue laboratory rabbit molecular cloning nucleic acid sequence platelets protein structure function tissue /cell culture transposon /insertion element virulence

项目摘要

DESCRIPTION (Adapted from applicants abstract): The direct binding of streptococci to human platelets is a postulated central mechanism in the pathogenesis of endocarditis. Bacterium-platelet binding may be critical for the attachment of blood-borne organisms to the valve surface, and for the subsequent formation of infected, macroscopic vegetations. The aim of this project is to define the molecular basis for the direct binding of Streptococcus sanguis to human platelets, and to determine the role of binding in the pathogenesis of endocarditis. By means of transposon mutagenesis, four isogenic mutants of Streptococcus sanguis strain M99 have been generated that bind platelets minimally in vitro. These mutants will provide a basis for the proposed research. A putative streptococcal gene ("spl~) encoding a ligand for human platelets will be identified, using probes derived from the low-binding mutants to screen a genomic library of strain M99, followed by cloning and sequencing. The spl gene product will then be purified, by cloning into a pET vector expression system. Once isolated, binding of the putative ligand to washed human platelets in vitro will be examined, using Scatchard analysis to determine if binding resembles a receptor-ligand interaction. The role of ligand mediated binding in the pathogenesis of endocarditis will be addressed in an animal model, by comparing the relative virulence of parental M99 with the isogenic mutant containing a Tn916deltaE insertion within the spl locus. By defining the mechanisms for streptococcal-platelet binding at the molecular level, this work will provide a basis for determining the role of platelets in the pathogenesis of endocarditis. In turn, this may provide a basis for developing novel diagnostic and therapeutic strategies.

描述（改编自申请人摘要）：直接约束力链球菌对人血小板的作用是假定的核心机制心内膜炎的发病机制。细菌-血小板结合可能至关重要用于将血源性生物附着到瓣膜表面，以及随后形成受感染的宏观植被。的目的该项目旨在定义直接结合的分子基础血链球菌对人血小板的作用并确定结合心内膜炎的发病机制。通过转座子诱变，血链球菌菌株 M99 的四个同基因突变体具有已在体外产生了最低限度地结合血小板的药物。这些突变体将为拟议的研究提供基础。假定的链球菌基因 (“spl~)编码人类血小板的配体将被鉴定，使用来自低结合突变体的探针用于筛选基因组文库菌株 M99，然后进行克隆和测序。 spl基因产物将然后通过克隆到 pET 载体表达系统中进行纯化。一次分离的，假定的配体在体外与洗涤的人血小板的结合将使用 Scatchard 分析进行检查，以确定结合是否类似于受体-配体相互作用。配体介导的结合的作用心内膜炎的发病机制将在动物模型中得到解决比较亲本 M99 与同基因突变体的相对毒力 spl 基因座内包含 Tn916deltaE 插入。通过定义链球菌-血小板在分子水平上的结合机制，这这项工作将为确定血小板在心内膜炎的发病机制。反过来，这可能会提供一个基础开发新的诊断和治疗策略。