RESEARCH OPPORTUNITY AWARD - CHARACTERIZATION OF NAG EXPRESSION AND FUNCTIONS

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目及 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 炭疽芽孢杆菌孢子如果释放到环境中，可导致严重的发病率和死亡率。需要有效的疫苗以及靶向免疫剂两者。目前获得许可的疫苗需要六次接种，每年加强一次，并与严重的不良事件有关。因此，已经做出了重大努力来开发改进的疫苗接种策略，其可以在减少疫苗接种的情况下产生持久的保护性免疫。詹姆斯博士的实验室一直致力于确定B的主要体液靶点。提供保护的炭疽菌。人类单克隆抗体以前是从接受炭疽疫苗的个体中产生的。这些单克隆抗体中的每一种先前已经在体外和体内毒素中和测定中以及通过线性表位和蛋白酶片段作图研究来表征。鉴定了多种功能信息性抗PA单克隆抗体。然而，这些功能信息单克隆抗体识别的结构表位仍不清楚。这些单克隆抗体显然识别构象表位而不是保护性抗原的线性肽表位。 本研究的目的是确定保护性抗原的结构决定簇，特异性识别的功能信息单克隆抗PA抗体。以炭疽PA的三维结构为指导，将PA的不同结构域克隆为重组融合蛋白。然后表达这些含有PA亚结构域的融合蛋白，并用于蛋白结合测定，以确定抗PA单克隆抗体结合所需的PA结构域。这些研究的结果将确定PA的哪些结构域是通过疫苗接种产生保护性免疫应答的最佳靶点。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Bacillus anthracis spores can cause significant morbidity and mortality if released into the environment. There is a need for both an effective vaccine as well as targeted immunotherapeutic agents. The currently licensed vaccine requires six vaccinations, annual boosters, and has been associated with serious adverse events. Thus there has been a major effort made to develop an improved vaccination strategy which can generate lasting protective immunity with reduced vaccinations. Dr. James' laboratory has been working to identify the major humoral targets of B. anthracis that provide protection. Human monoclonal antibodies were previously generated from individuals who received the anthrax vaccine. Each of these monoclonal antibodies has previously been characterized in both in vitro and in vivo toxin neutralization assays as well as by linear epitope and protease fragment mapping studies. Multiple functionally informative anti-PA monoclonal antibodies were identified. However, the structural epitopes recognized by these functionally informative monoclonal antibodies remained unclear. These monoclonal antibodies apparently recognized conformational epitopes rather than the linear peptide epitopes of protective antigen. The goal of this study was to define the protective antigen structural determinants that were specifically recognized by the functionally informative monoclonal anti-PA antibodies. Using the three dimensional structure of anthrax PA as a guide, different domains of PA will be cloned as recombinant fusion proteins. These PA subdomain containing fusion proteins will then be expressed and used in protein binding assays to define which PA domains are required for anti-PA monoclonal antibody binding. The results of these studies will identify which domains of PA are the best targets for generating protective immune responses through vaccination.