Self-association motifs in the enteroaggregative Escherichia coli heat-resistant agglutinin 1
Self-association motifs in the enteroaggregative Escherichia coli heat-resistant agglutinin 1
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DOI：
10.1099/mic.0.000303
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发表时间：
2016-07-01
期刊：
影响因子：
2.8
通讯作者：
Okeke, Iruka N.
中科院分区：
文献类型：
--
作者：
Glaubman, Jessica;Hofmann, Jennifer;Okeke, Iruka N.
耐热凝集素1（Hra 1）是一种完整的外膜蛋白，存在于大肠杆菌菌株中，这些菌株是特殊的殖民者。Hra 1来自肠聚集性E.大肠杆菌菌株042足以赋予对人上皮细胞的粘附并引起细菌自身聚集。Hra 1与Tia侵袭素密切相关，Tia侵袭素也赋予粘附性，但不赋予自动聚集性。在这里，我们已经证明，Hra 1介导的自聚集的自我协会，我们假设，至少有一些表面暴露的氨基酸序列是存在于Hra 1，但缺乏在Tia，代表自聚集基序。我们将FLAG标签沿着Hra 1的长度插入，并使用免疫斑点印迹来验证四个计算机预测的外环确实是表面暴露的。在Hra 1中，我们将其中三个环中的九个候选基序（长度从一到十个氨基酸不等）交换到Tia中的相应序列。其中三个基序是Hra 1介导的自聚集所必需的。在数据库中搜索含有这些基序的其他表面蛋白; GGXWRDDXK基序也存在于Rck的表面暴露区域，Rck是一种沙门氏菌血清型鼠伤寒补体抗性蛋白。克隆和位点特异性突变表明，Rck可以赋予弱，GGXWRDDXK依赖性自聚集的自我协会。Hra 1和Rck似乎形成异源缔合，并且Hra 1-Rck缔合在两种分子上都需要GGXWRDDXK。然而，GGYWRDDLKE肽不足以干扰Hra 1介导的自聚集。在本研究中，已确定了三个自聚集基序在一个完整的外膜蛋白，它被证明，其中至少有一个工作在不同的细胞表面的上下文中。
The heat-resistant agglutinin 1 (Hra1) is an integral outer membrane protein found in strains of Escherichia coli that are exceptional colonizers. Hra1 from enteroaggregative E. coli strain 042 is sufficient to confer adherence to human epithelial cells and to cause bacterial autoaggregation. Hra1 is closely related to the Tia invasin, which also confers adherence, but not autoaggregation. Here, we have demonstrated that Hra1 mediates autoaggregation by self-association and we hypothesize that at least some surface-exposed amino acid sequences that are present in Hra1, but absent in Tia, represent autoaggregation motifs. We inserted FLAG tags along the length of Hra1 and used immune-dot blots to verify that four in silico-predicted outer loops were indeed surface exposed. In Hra1 we swapped nine candidate motifs in three of these loops, ranging from one to ten amino acids in length, to the corresponding sequences in Tia. Three of the motifs were required for Hra1-mediated autoaggregation. The database was searched for other surface proteins containing these motifs; the GGXWRDDXK motif was also present in a surface exposed region of Rck, a Salmonella enterica serotype Typhimurium complement resistance protein. Cloning and site-specific mutagenesis demonstrated that Rck can confer weak, GGXWRDDXK-dependent autoaggregation by self-association. Hra1 and Rck appear to form heterologous associations and GGXWRDDXK is required on both molecules for Hra1-Rck association. However, a GGYWRDDLKE peptide was not sufficient to interfere with Hra1-mediated autoaggregation. In the present study, three autoaggregation motifs in an integral outer membrane protein have been identified and it was demonstrated that at least one of them works in the context of a different cell surface.