Mapping critical interactive sites within the periplasmic domain of the Vibrio cholerae type II secretion protein EpsM
Mapping critical interactive sites within the periplasmic domain of the Vibrio cholerae type II secretion protein EpsM
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DOI：
10.1128/jb.01256-07
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发表时间：
2007-12-01
影响因子：
3.2
通讯作者：
Sandkvist, Maria
中科院分区：
文献类型：
--
作者：
Johnson, Tanya L.;Scott, Maria E.;Sandkvist, Maria
II型分泌物(T2S)系统存在于许多革兰氏阴性菌中，包括致病和非致病物种，它支持各种毒素、蛋白酶和脂肪酶进入细胞外环境。在霍乱弧菌中，T2S装置由12个EPS蛋白组成，这些蛋白组装成一个跨越整个细胞膜的多蛋白复合体。其中两种蛋白质，EpsM和EPSL，是存在于内膜的分泌机制的关键组件。除了可能形成同源二聚体外，EPSL和EpsM还被证明在内膜形成稳定的复合体，并相互保护对方免受蛋白质降解的影响。为了确定和定位与EpsM和EPSL的另一个分子参与蛋白质-蛋白质相互作用的EpsM的特定区域，我们通过功能鉴定、共沉淀和免疫共沉淀测试了EpsM的缺失结构与全长EpsM和EPSL的相互作用。对截短的EpsM突变体的分析表明，EpsM在氨基酸100到135之间的区域是形成同源低聚物所必需的，而残基84到99似乎是与EPSL稳定相互作用的关键。
The type II secretion (T2S) system is present in many gram-negative species, both pathogenic and nonpathogenic, where it supports the delivery of a variety of toxins, proteases, and lipases into the extracellular environment. In Vibrio cholerae, the T2S apparatus is composed of 12 Eps proteins that assemble into a multiprotein complex that spans the entire cell envelope. Two of these proteins, EpsM and EpsL, are key components of the secretion machinery present in the inner membrane. In addition to likely forming homodimers, EpsL and EpsM have been shown to form a stable complex in the inner membrane and to protect each other from proteolytic degradation. To identify and map the specific regions of EpsM involved in protein-protein interactions with both another molecule of EpsM and EpsL, we tested the interactions of deletion constructs of EpsM with full-length EpsM and EpsL by functional characterization and copurification as well as coimmunoprecipitation. Analysis of the truncated EpsM mutants revealed that the region of EpsM from amino acids 100 to 135 is necessary for EpsM to form homo-oligomers, while residues 84 to 99 appear to be critical for a stable interaction with EpsL.