GENETIC ANALYSIS OF CHOLERA TOXIN STRUCTURE AND FUNCTION

描述（改编自申请人摘要）：霍乱是一种大流行病 由霍乱弧菌引起的疾病，其中霍乱毒素（CT）引起 危及生命的腹泻 产肠毒素大肠大肠杆菌和其他肠道细菌 肠道细菌引起与霍乱密切相关的腹泻病。 高度 目前还没有针对这些疾病的有效疫苗。 我们漫长 范围目标是了解霍乱毒素的结构和功能 (CT)在分子水平上， 全毒素病 具体目标1将使用生化，遗传，细胞 生物学和结构学方法来表征CT-A和CT-B的特征 决定CT的生物活性。 这些研究将分析 CT-A和CT-B之间的相互作用对于CT的组装是必不可少的， 确定CT-B的构象依赖性表位的结构， 中和抗体，研究CT-A的活化途径， 确定活性CT-A1的结构，探讨其特异性作用 神经节苷脂在CT和相关肠毒素的细胞内运输中， 并开发CT-A跨细胞内膜易位的检测方法。 具体目标2将评估全毒素样嵌合体，其中微生物 保护性抗原替代CT的A1结构域，作为模型口腔因子， 将测试霍乱弧菌诱导保护性抗毒性的能力， 和抗霍乱的抗菌免疫力;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， 多种荚膜血清型肺炎球菌。

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.