Vesicular stabilization and activity augmentation of enterohaemorrhagic Escherichia coli haemolysin
Vesicular stabilization and activity augmentation of enterohaemorrhagic Escherichia coli haemolysin
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DOI：
10.1111/j.1365-2958.2009.06618.x
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发表时间：
2009-03-01
影响因子：
3.6
通讯作者：
Karch, Helge
中科院分区：
文献类型：
--
作者：
Aldick, Thomas;Bielaszewska, Martina;Karch, Helge
肠出血性大肠杆菌溶血素(EHEC-HLY)属于毒素重复序列(Repeat-in-Toxin，RTX)家族，其成员通过自身聚集迅速灭活自身，是EHEC的一个潜在毒力因子。通过研究EHEC-HLY的胞外分泌状态，我们发现该毒素既以游离的、可溶的形式存在，又与EHEC挤出的外膜囊泡(OMV)有很高的结合倾向，且与其酰化状态无关。我们用扫描电子显微镜和结合实验比较了这两种毒素形式与红细胞的相互作用。OMV相关毒素在生理条件下的稳定性比游离EHEC-HLY高80倍，溶血活性延长(半衰期为20h比15min)。在溶血之前，携带EHEC-HLY的OMV与红细胞的钙依赖结合；这种结合是由EHEC-HLY介导的。我们证明EHEC-HLY在OMV中是一种具有生物活性的货物，具有双重作用：细胞结合蛋白和溶血素。这些成对的功能产生了一种与OMV相关的RTX毒素的生物有效形式，并增强了它对靶细胞的潜力。我们的发现为RTX毒素的稳定提供了一个总体概念，并为这些重要毒力因子的生物学提供了新的见解。
Haemolysin from enterohaemorrhagic Escherichia coli (EHEC-Hly), a putative EHEC virulence factor, belongs to the RTX (repeat-in-toxin) family whose members rapidly inactivate themselves by self-aggregation. By investigating the status of EHEC-Hly secreted extracellularly, we found the toxin both in a free, soluble form and associated, with high tendency and independently of its acylation status, to outer membrane vesicles (OMVs) extruded by EHEC. We compared the interaction of both toxin forms with erythrocytes using scanning electron microscopy and binding assays. The OMV-associated toxin was substantially (80 times) more stable under physiological conditions than the free EHEC-Hly as demonstrated by prolonged haemolytic activity (half-life time 20 h versus 15 min). The haemolysis was preceded by calcium-dependent binding of OMVs carrying EHEC-Hly to erythrocytes; this binding was mediated by EHEC-Hly. We demonstrate that EHEC-Hly is a biologically active cargo in OMVs with dual roles: a cell-binding protein and a haemolysin. These paired functions produce a biologically potent form of the OMV-associated RTX toxin and augment its potential towards target cells. Our findings provide a general concept for stabilization of RTX toxins and open new insights into the biology of these important virulence factors.