StcE, an E.coli O157:H7 Protease Specific for C1-Inh

StcE，一种针对 C1-Inh 的大肠杆菌 O157:H7 蛋白酶

• 批准号: 6579204
• 负责人: Rodney A. Welch
• 金额: $ 28.62万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-01-16 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6579204
• 关键词: Escherichia coli 0157:H7; SDS polyacrylamide gel electrophoresis; bacteria infection mechanism; bacterial cytopathogenic effect; bacterial proteins; clinical research; densitometry; diarrhea; endopeptidases; enzyme linked immunosorbent assay; feces analysis; gastrointestinal epithelium; gene expression; human subject; immunoelectroadsorption; laboratory rabbit; pathologic process; protein binding; protein sequence; radioimmunoassay; shiga toxin; swine; western blottings

DESCRIPTION (Provided by applicant): Enterohemorrhagic Escherichia coli (EHEC), principally serotype O157:H7, cause an estimated 20,000 cases of diarrheal disease in the United States per year. 2-6 percent of the infected individuals, mostly young children progress to a severe renal disease, hemolytic uremic syndrome (HUS). The EHEC pathogenic factors that lead to bloody colitis and HUS are poorly understood, but knowledge of some mechanisms has recently emerged. Intimin-mediated adherence and type III effectors are encoded by a chromosomal locus termed LEE. The phage-encoded Shiga toxins (Stxs) are responsible for significant aspects of EHEC disease. EHEC strains commonly possess large plasmids, the prototype being pO157. We have identified a new pO157 gene, stcE, which encodes an extracellular zinc-metalloendoprotease (ZMP) that specifically cleaves the critical anti-inflammatory regulator C l-esterase inhibitor (C 1-Inh). C 1-Inh is a serine protease inhibitor (serpin) that provides the principal inhibition of the proteolytic cascades involved in classic and mannan-binding ligand complement activation, contact activation and intrinsic coagulation. C l-Inh inhibits diverse proteases: Clr and Cls, MASP-1, MASP-2, kallikrein, FXIIa, FXIa, and plasmin. Deficiencies in Cl-Inh cause profound clinical syndromes. The best known is hereditary angioedema (HAE), a genetic deficiency in Cl-Inh, which is characterized by transient, recurrent attacks of intestinal cramps, vomiting, diarrhea and life-threatening episodes of tracheal swelling. Fluorescenated StcE binds to cultured macrophages, B- and T-cells. Thus, StcE is an example of a growing class of ZMPs such as tetanus, botulinum and anthrax lethal factor toxins. These ZMPs, in contrast to the homologous Pseudomonas and Vibrio ZMPs, have specific, non-extracellular matrix protein targets. We will test the hypothesis that StcE degrades soluble or cell-associated Cl-Inh, and this results in poorly regulated serine protease cascades involving complement activation, contact activation and coagulation. This dysregulation would then contribute to local inflammation, tissue damage and edema. The elucidation of StcE structure and function(s) may result in new targets for chemotherapeutic or immune prevention or treatment of EHEC infections, which now are best managed only by supportive therapy.

描述（由申请人提供）：肠出血性大肠杆菌（EHEC），主要是血清型O157:H7，在美国每年导致约20,000例腹泻病例。2- 6%的感染者，主要是幼儿，发展为严重的肾脏疾病，溶血性尿毒症综合征（HUS）。导致血性结肠炎和溶血性尿毒综合征的肠出血性大肠杆菌致病因素尚不清楚，但最近出现了一些机制的知识。抗体介导的粘附和III型效应物是由染色体位点LEE编码的。噬菌体编码的志贺毒素（Stxs）是肠出血性大肠杆菌疾病的重要原因。肠出血性大肠杆菌菌株通常具有较大的质粒，原型为pO157。我们发现了一个新的pO157基因，stcE，它编码细胞外锌-金属内源性蛋白酶（ZMP），特异性地切割关键的抗炎调节剂C -酯酶抑制剂（c1 - inh）。c1 - inh是一种丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶），对经典和甘露聚糖结合配体补体激活、接触激活和内在凝血过程中涉及的蛋白水解级联反应提供主要抑制作用。C l-Inh抑制多种蛋白酶：Clr和Cls、MASP-1、MASP-2、钾化因子、FXIIa、FXIa和纤溶蛋白。缺乏Cl-Inh会引起严重的临床症状。最著名的是遗传性血管性水肿（HAE），这是一种Cl-Inh基因缺陷，其特征是短暂的、反复发作的肠痉挛、呕吐、腹泻和危及生命的气管肿胀发作。荧光StcE与培养的巨噬细胞、B细胞和t细胞结合。因此，StcE是破伤风、肉毒杆菌和炭疽致死因子毒素等一类日益增多的zmp的一个例子。与同源假单胞菌和弧菌相比，这些ZMPs具有特异性的非细胞外基质蛋白靶点。我们将验证StcE降解可溶性或细胞相关Cl-Inh的假设，这导致包括补体激活、接触激活和凝固在内的调节不良的丝氨酸蛋白酶级联反应。这种失调会导致局部炎症、组织损伤和水肿。StcE结构和功能的阐明可能为肠出血性大肠杆菌感染的化疗或免疫预防或治疗提供新的靶点，目前只有通过支持治疗才能最好地管理这些感染。