NEUTRALIZING EPITOPES OF CHLAMYDIA TRACHOMATIS

沙眼衣原体的中和表位

• 批准号: 3145486
• 负责人: ELLENA Marie PETERSON
• 金额: $ 16.67万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1994-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3145486
• 关键词: Chlamydia trachomatis; active immunization; antibody neutralization test; bacterial antigens; complement; crosslink; enzyme linked immunosorbent assay; epitope mapping; laboratory mouse; membrane proteins; monoclonal antibody; neutralizing antibody; passive immunization; protein structure function; serum; synthetic peptide

The goal of this proposal is to identify and characterize continuous neutralizing epitopes of the major outer membrane protein (MOMP) of Chlamydia trachomatis. This organism is the leading cause of preventable blindness and sexually transmitted diseases, therefore protection through an effective vaccine would be desirable. Vaccine trials in the past which have employed the whole organism for immunization have failed, partly due to hypersensitivity reactions. An alternate approach to immunization is to engineer a subunit vaccine where only epitopes that elicit a protective host response are included. To identify such key epitopes MOMP was chosen as the focus of this investigation since it is surface exposed, has been shown to elicit neutralizing antibodies and DNA sequence data is available and thus the derived amino add sequence. More specifically variable domain (VD) IV will be the main focus of this proposal since antibodies that neutralize the B-,B-related and C-related complexes have been mapped to this region. However, MAbs that have mapped to this region fail to neutralize members of the C-complex Therefore, in addition to VD IV, VD I and VD 11 will be examined with C-complex serovars in order to be more successful in identifying continuous neutralizing epitopes that are broadly reactive and neutralize these members of the species. Peptides representing VD IV from serovars L2 and E and VD I, II and IV from serovar C will be used to immunize mice. Both the polyclonal sera as well as MAbs resulting from the immunizations will be examined for their ability to neutralize C. trachomatis. This will be accomplished by performing in vitro neutralization assays. In addition, the Geysen technique will be used to map recognition sites by ELISA using overlapping hexameric peptides to the VD used in the immunizations. The functional, i.e., neutralizable, site will be determined by competition experiments in which the neutralizing antibodies will be competed for by viable C. trachomatis and peptides to the region of interest. The spectrum of serovars neutralized by the antibodies will be established as well as the ability of the antibodies to act in an additive or synergistic manner. Mice will be immunized with peptide conjugates or passively immunized with MAbs that are identified as broadly neutralizing in terms of serovars. These animals will then be challenged with C. trachomatis in both a systemic as well as a mucosa] model of infection. In addition, studies aimed at determining the mechanism(s) of inhibition of the broad neutralizing MAbs will be performed. The work proposed in this investigation will provide structure-function information about a key structural protein of this organism and will be useful in the future design of vaccines to control the spread and growth of this important human pathogen.

该提案目标是识别和描述连续的 猪瘟病毒主要外膜蛋白(MOMP)中和表位 沙眼衣原体。这种微生物是可预防疾病的主要原因 失明和性传播疾病，因此通过 一种有效的疫苗将是可取的。过去的疫苗试验 利用整个生物体进行免疫的方法都失败了， 部分原因是过敏反应。另一种方法是 免疫就是设计一种亚单位疫苗，在这种疫苗中只有表位 诱导保护性宿主反应也包括在内。来识别这样的密钥 表位MOMP被选为本次研究的重点，因为它是 表面暴露，已被证明可引发中和抗体和DNA 序列数据是可用的，因此衍生的氨基添加序列。更多 具体来说，可变区(Vd)IV将是这一研究的主要焦点 建议自中和B-、B-相关和C-相关抗体以来 复合体已经被绘制到这个地区。然而，已经拥有的单抗 映射到这个区域不能中和C-复合体的成员 因此，除VDIV、VIDI和VD11外，还将用 C-复合体血清型，以便更成功地识别 具有广泛反应性和中和性的连续中和表位 这些物种的成员。代表血清型VD IV的多肽 将使用C血清型的L2和E以及VD I、II和IV进行免疫 老鼠。产生的多克隆血清和单抗 将检查免疫接种是否具有中和C. 沙眼衣原体。这将通过体外实验来完成。 中和试验。此外，盖森技术将被用于 利用重叠六聚体多肽的酶联免疫吸附试验绘制识别位点图 免疫中使用的VD。官能化的，即可中和的， 选址将由竞赛实验确定，在这些实验中， 中和抗体将被存活的沙眼衣原体和 将多肽转移到感兴趣区域。血清型的光谱被中和了 通过抗体的建立以及抗体的能力 抗体以相加或协同的方式起作用。老鼠会成为 用多肽结合物免疫或用以下单抗被动免疫 在血清型方面被鉴定为广泛中和。这些 然后，动物将在两种系统性AS中被沙眼衣原体挑战 以及粘膜]感染的模型。此外，针对以下方面的研究 广谱中和单抗抑制机理的确定(S) 将会被执行。这项调查中提出的工作将提供 一种关键结构蛋白的结构-功能信息 生物体，将在未来的疫苗设计中有用，以控制 这种重要的人类病原体的传播和生长。