Genetic and biochemical study ov V. cholerase RTX toxin

V.霍乱酶 RTX 毒素的遗传和生化研究

基本信息

批准号：
7152560
负责人：
Karla J F Satchell
金额：
$ 28.16万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-12-15 至 2008-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7152560
关键词：
Accounting Actin-Binding Protein Actins Acyltransferase Address Affect Amino Acids Animal Model Applications Grants Attenuated Live Virus Vaccine Bacterial Toxins Binding Biochemical Biochemical Genetics Biological Biological Assay C-terminal Catalytic Domain Cell Fractionation Cell Line Cells Cholera Cholera Toxin Chromosomes Computer Simulation Computers Cytoplasm Data Defect Diarrhea Disease Environment Extravasation F-Actin Factor V Family Fibrinogen Future G Actin Genes Genetic High Pressure Liquid Chromatography Human Volunteers In Vitro Inflammatory Intestines Investigation Life Link Liquid substance Maintenance Membrane Methods Microscopy N-terminal Nature Organism Pathogenesis Pathway interactions Post-Translational Protein Processing Production Property Proteins Proteolytic Processing Reaction Reagent Research Resiniferatoxin Role Stress Fibers Structure Symptoms Testing Toxin Ultrafiltration Vibrio Vibrio cholerae Virulence Virulence Factors base crosslink cytotoxic cytotoxicity deletion analysis depolymerization dimer in vivo insight interest member mutant novel pathogen polarized cell polypeptide response size tissue/cell culture toxin V

项目摘要

Vibrio cholerae strains that do not produce cholera toxin induce a more inflammatory diarrhea than normal cholera disease, implicating other potent toxins in the pathogenesis of cholera Several accessory toxins of V cholerae are purported to account for this reactogenic response One of these factors is the newly discovered VcRtxA toxin of V cholerae VcRtxA is a large protein toxin that is a unique member of the RTX family Production of this toxin has been evolutionarily conserved by Vibrio sp indicating that maintenance of this large toxin is essential for virulence or survival in the environment Its cytotoxicity has been further shown to function by a novel mechanism This toxin causes depolymerization of actin stress fibers in both polarized and non-polarized cell lines by a unique pathway Concurrent with depolymerization, the actin molecules become covalently linked together into dimers, trimers, and higher order multimers This observation suggests that covalent crosslinking of actin by the toxin drives actin depolymerization This unusual reaction distinguishes VcRtxA from all other bacterial toxins that cause actin depolymerization Research performed under this grant proposal will investigate further the novel biochemical properties of this important virulence factor The VcRtxA toxin is the largest single polypeptide protein toxin ever described, however, it is unclear whether the full 4545 amino acid protein is the toxic moiety The size of the toxic moiety and potential post-translational modifications will by identified by examining biochemical properties of the active form of purified toxin The catalytic activity will then be investigated and the role of a putative catalytic domain in this reaction will be assessed tt will then be established whether the toxin is active at the membrane or within the cytoplasm of the target cells, and the role of a putative actin binding domain will be characterized

不产生霍乱毒素的弧菌霍乱菌株会诱发比炎症性腹泻的正常的霍乱疾病，暗示其他有效的毒素在霍乱的发病机理中 V霍乱的V毒素据称是为了解释这种反应的反应，其中一个因素之一是新发现的V霍乱VCRTXA的VCRTXA毒素是一种大蛋白毒素，是独特的成员这种毒素的RTX家族产生在进化上是由纤维中的SP进化保存的，表明维持这种大毒素对于在其细胞毒性的环境中的毒力或生存至关重要进一步证明了这种毒素引起肌动蛋白应激的解聚作用的作用通过与解聚的独特途径同时发生的纤维和非极化细胞系中的纤维，肌动蛋白分子将共价链接到二聚体，三聚体和高阶多聚体中观察表明，毒素驱动肌动蛋白解聚的肌动蛋白的共价交联异常反应将VCRTXA与引起肌动蛋白解聚的所有其他细菌毒素区分开根据该赠款提案进行的研究将进一步研究此的新型生化特性重要的毒力因子VCRTXA毒素是有史以来描述的最大的单多肽蛋白毒素，但是，目前尚不清楚整个4545氨基酸蛋白是否是有毒的有毒部分部分通过检查的生化特性，将识别然后，将研究纯化毒素的活性形式，然后研究催化活性并提出推定的作用该反应中的催化结构域将被评估，然后将确定毒素是否有效在膜或靶细胞的细胞质内，推定肌动蛋白结合结构域的作用将为特征