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，一直受到阻碍，无法 成功地在体外高产量培养螺旋体， 持续的时间。 虽然T.苍白球可以是 在兔子睾丸中繁殖并少量纯化， 该方法通常导致密螺旋体毒力的丧失。 最近，一些研究人员试图使用重组 DNA技术试图绕过一些 获得合适数量的T. 用于实验研究的苍白球组分。 本实验室 我们正试图在大肠杆菌中克隆和表达， 编码T.苍白球细胞表面和 细胞外蛋白抗原 我们特别感兴趣的是 这些蛋白质，因为它们是唯一的密螺旋体蛋白， 与受感染的宿主直接接触， 在发病机制中的重要作用。 据推测，一种保护性免疫 免疫应答将针对这些抗原。 我们的长期 目标是产生细胞表面和细胞外蛋白 足够数量的抗原，以便它们在免疫系统中的作用。 梅毒的发病机制可以调查和他们的适用性 用作改进的血清诊断测试试剂和/或潜在的 可以评估疫苗原。 在我们的项目期间，我们建议做以下事情： (i)鼠单克隆抗体和兔多克隆抗体 与栽培苍白球细胞表面和细胞外蛋白抗原 将被生成。 (ii)E.表达目的蛋白抗原的大肠杆菌克隆将 和战略，以最大限度地表达这些 E.大肠杆菌将被开发，从而促进其 洁净. 我们手头上有一个T.苍白球基因组 使用质粒载体构建的DNA。 我们有 开发了鉴定和表征E.大肠杆菌克隆 表达密螺旋体蛋白抗原，并产生 足够量的蛋白质，允许纯化。 (iii)各种旨在确定生物学的实验 细胞表面和细胞外T. 螺旋体 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.