Identification of Sat, an autotransporter toxin produced by uropathogenic Escherichia coli
Identification of Sat, an autotransporter toxin produced by uropathogenic Escherichia coli
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DOI：
10.1046/j.1365-2958.2000.02110.x
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发表时间：
2000-10-01
影响因子：
3.6
通讯作者：
Mobley, HLT
中科院分区：
文献类型：
--
作者：
Guyer, DM;Henderson, IR;Mobley, HLT
尿路感染(UTI)是一种非常常见的肠外感染，而大肠埃希菌是迄今为止最常见的致病微生物。致尿性大肠杆菌具有不同于共生菌株的特征，例如允许毒力因子靶向胞外隔室的分泌系统。至少五种特征性的分泌机制之一是自动转运蛋白系统，它涉及蛋白质通过内膜转运，推测是通过SEC系统，并通过由羧基末端自转运蛋白结构域形成的β-桶状孔蛋白结构穿过外膜。我们鉴定出一个107 kDa的蛋白在与急性肾盂肾炎临床综合征相关的大肠杆菌菌株中的表达频率显著高于粪便菌株(P=0.029)。我们从一名急性肾盂肾炎患者的血液和尿液培养的E.ColiCFT073中分离出这种蛋白。N端氨基酸序列与已知的两个SpATE(肠杆菌科丝氨酸蛋白酶自身转运蛋白)蛋白Pet和ESPC具有最高的相似性，利用来自pET 5‘区的509bp的探针，从大肠杆菌CFT073基因文库中筛选出10个粘粒克隆进行杂交。从一个粘粒克隆中，发现了一个7.5kb的EcoRI限制性片段，该片段与探针反应强烈，包含了整个3885bp的基因。预测的142 kDa蛋白产物具有Spate自身转运蛋白的三个典型结构域：一个由49个氨基酸组成的异常长的信号序列；一个107 kDa的乘客区，包含一个共同的丝氨酸蛋白酶活性位点(GDSGSG)；以及一个C端的自转运蛋白结构域，其大小为30 kDa。该蛋白在Vero原代肾、HK-2膀胱和Hep-2细胞上具有丝氨酸蛋白酶活性和细胞病变活性，其名称Sat(分泌型自体转运蛋白毒素)就是从这些特性而来的。此外，在感染E.ColiCFT073的小鼠血清中存在Sat抗体，根据其与致病菌株的相关性、其细胞病变表型以及感染后激发强烈抗体反应的能力，我们推测Sat代表了一种新的尿路致病性E-Coli的毒力决定因素。
Urinary tract infection (UTI) is a very common extraintestinal infection, and Escherichia coli is by far the most common causative organism. Uropathogenic E. coli possess traits that distinguish them from commensal strains of E, coli, such as secretion systems that allow virulence factors to be targeted to extracytoplasmic compartments. One of at least five characterized secretion mechanisms is the autotransporter system, which involves translocation of a protein across the inner membrane, presumably via the sec system, and across the outer membrane through a beta-barrel porin structure formed by the carboxy-terminus autotransporter domain. We identified a 107 kDa protein that was expressed significantly more often by E. coli strains associated with the clinical syndrome of acute pyelonephritis than by faecal strains (P = 0.029). We isolated the protein from E. coli CFT073, a strain cultured from the blood and urine of a patient with acute pyelonephritis. The N-terminal amino acid sequence showed highest similarity to two known SPATE (serine protease autotransporters of Enterobacteriaceae) proteins, Pet and EspC, Using a 509 bp probe from the 5' region of pet, 10 cosmid clones of an E. coli CFT073 gene library were positive for hybridization. From one cosmid clone, a 7.5 kb EcoRI restriction fragment, which reacted strongly with the probe, was shown to include the entire 3885 bp gene. The predicted 142 kDa protein product possesses the three domains that are typical of SPATE autotransporters: an unusually long signal sequence of 49 amino acids; a 107 kDa passenger domain containing a consensus serine protease active site (GDSGSG); and a C-terminal autotransporter domain of 30 kDa. The protein exhibited serine protease activity and displayed cytopathic activity on VERO primary kidney, HK-2 bladder and HEp-2 cell lines; the name Sat (Secreted autotransporter toxin) was derived from these properties. In addition, Sat antibodies were present in the serum of mice infected with E. coli CFT073, Based upon its association with pathogenic isolates, its cytopathic phenotype and its ability to elicit a strong antibody response after infection, we postulate that Sat represents a novel Virulence determinant of uropathogenic E, coli.