SURFACE & EXTRACELLULAR ANTIGENS OF T PALLIDUM

梅毒是一种传染性的性传播疾病 仍然是美国的一个主要公共卫生问题 以及其他国家。梅毒螺旋体的研究 梅毒的病原体，一直受到无法 成功地将螺旋体在体外进行了高产培养 持续的时间段。尽管梅毒螺旋体可以 在兔睾丸中繁殖并少量纯化，如 手术通常会导致梅毒螺旋体毒性丧失。 最近，几名调查人员试图使用重组 DNA技术试图绕过一些 获得适当数量的T。 用于实验研究的梅毒成分。在我们的实验室 我们正试图克隆并在大肠杆菌中表达 梅毒螺旋体细胞表面编码的遗传信息和 胞外蛋白抗原。我们特别感兴趣的是 因为它们是唯一的梅毒螺旋体蛋白 与受感染的主机直接接触，因此可能会扮演 在发病机制中起重要作用。据推测，一种保护性免疫 应对措施将针对这些抗原。我们的长期计划 目标是产生细胞表面和胞外蛋白 足够数量的抗原，以使它们在 可以研究梅毒的发病机制及其适用性 用作改进的血清诊断检测试剂和/或潜在的 接种疫苗的人可以被评估。 在我们的项目期内，我们建议做以下工作： (I)鼠单抗和兔多克隆抗体 对梅毒螺旋体细胞表面和胞外蛋白抗原的影响 将会被生成。 (Ii)表达目的蛋白抗原的大肠杆菌克隆将 确定并制定策略，以最大限度地表达这些 大肠杆菌中的蛋白质将被开发出来，从而促进其 净化。我们手头有一个梅毒螺旋体基因组文库 使用质粒载体构建的DNA。我们有 已开发的鉴定和鉴定大肠杆菌克隆的技术 表达梅毒螺旋体蛋白抗原并产生 足够数量的蛋白质，可以进行提纯。 (Iii)旨在确定生物量的各种实验 梅毒螺旋体细胞表面和胞外的意义 将进行蛋白质抗原。

Syphilis is an infectious, sexually transmitted disease that continues to be a major public health problem in the United States as well as other countries. Research on Treponema pallidum, the etiological agent of syphilis, has been hampered by the inability to successfully culture the spirochete to high yields in vitro for sustained periods of time. Although T. pallidum can be propagated in rabbit testes and purified in small quantities such procedures usually result in the loss of treponemal virulence. Recently, several investigators have sought to use recombinant DNA technology in an attempt to circumvent some of the difficulties associated with obtaining suitable quantities of T. pallidum components for experimental studies. In our laboratory we are attempting to clone and express in Escherichia coli and the genetic information encoding for T. pallidum cell surface and extracellular protein antigens. We are particularly interested in these proteins since they are the only treponemal proteins that are in direct contact with the infected host and thus may play an important role in pathogenesis. Presumably, a protective immune response would be targeted against such antigens. Our long term goal is to produce the cell surface and extracellular protein antigens in sufficient quantities so that their role in the pathogenesis of syphilis can be investigated and their applicability for use as improved serodiagnostic test reagents and/or potential vaccinogens can be evaluated. For our project period, we propose to do the following: (i) Murine monoclonal antibodies and rabbit polyclonal antibodies to the T. pallidum cell surface and extracellular protein antigens will be generated. (ii) E. coli clones expressing the protein antigens of interest will be identified and strategies to maximize expression of these proteins in E. coli will be developed, thus facilitating their purification. We have on hand a library of T. pallidum genomic DNA that was constructed using a plasmid vector. We have developed techniques to identify and characterize E. coli clones expressing treponemal protein antigens and to produce the proteins in sufficient quantities that allowed for purification. (iii) a variety of experiments aimed at determining the biological significance of the cell surface and extracellular T. pallidum protein antigens will be conducted.