GENETIC ANALYSIS OF CHOLERA TOXIN STRUCTURE AND FUNCTION

描述(摘自申请者的摘要)：霍乱是一种流行病 霍乱弧菌引起的疾病，霍乱毒素(CT)引起 危及生命的腹泻。产肠毒素大肠埃希菌和其他肠毒素 肠道细菌引起与霍乱密切相关的腹泻疾病。高度 目前还没有针对这些疾病的有效疫苗。我们的龙 范围目标是了解霍乱毒素的结构和功能 (CT)在分子水平上，并利用其显著的性质 全毒素疾病。特殊目标1将使用生化、遗传、细胞 表征CT-A和CT-B特征的生物学和结构方法 它们决定了CT的生物活性。这些研究将分析 CT-A和CT-B之间的相互作用是CT组装所必需的， 确定CT-B的构象依赖表位的结构 中和抗体，研究CT-A和CTA的激活途径 确定活性CT-A1的结构，探索其特异性作用 神经节苷脂在CT和相关肠毒素的细胞内转运中的作用 并开发CT-A跨细胞内膜转运的方法。 特定目标2将评估类全息毒素嵌合体，其中微生物 保护性抗原取代CT的A1结构域，成为模范口腔因子 霍乱弧菌将接受诱导保护性抗毒素能力测试 和对霍乱的抗菌免疫；以及融合了 肺炎链球菌PSPA表达保守的保护性表位， 将测试它们诱导交叉保护免疫的能力 多种包膜血清型的肺炎球菌。

DESCRIPTION (Adapted from applicant's abstract): Cholea is a pandemic disease caused by V. cholerae in which cholera toxin (CT) cause life-threatening diarrhea. Enterotoxigenic E. coli and other enteroxic enteric bacteria cause diarrheal disease closely related to cholera. Highly effective vaccines against these diseases are not yet available. Our long range goals are to understand the structure and function of cholera toxin (CT) at the molecular level and to exploit the remarkable properties of holotoxin diseases. Specific Aim 1 will use biochemical, genetic, cell biological, and structural methods to characterize features of CT-A and CT-B that determine the biological actives of CT. These studies will analyze interactions between CT-A and CT-B that are essential for assembly of CT, determine structure of conformation-dependent epitopes of CT-B that elicit neutralizing antibodies, investigate the activation pathway for CT-A and determine the structure of active CT-A1, explore the role of specific gangliosides in intracellular trafficking of CT and related enterotoxins, and develop assays for translocation of CT-A across intracellular membranes. Specific Aim 2 will evaluate holotxin-like chimeras, in which microbial protective antigens replace the A1 domain of CT, as model oral factors of Vibrio Cholerae, will be tested for ability to induce protective anti-toxic and anti-bacterial immunity against cholera; and chimeras that incorporate Streptococcus pneumoniae PspA, which express conserved protective epitopes, will be tested for their ability to induce cross-protective immunity against pneumococci of multiple capsular serotypes.