Structural and Functional Analysis of the Escherichia coli Acid-Sensing Histidine Kinase EvgS.
Structural and Functional Analysis of the Escherichia coli Acid-Sensing Histidine Kinase EvgS.
复制标题
大肠杆菌酸性组氨酸激酶EVG的结构和功能分析。
DOI：
10.1128/jb.00310-17
复制
发表时间：
2017-09-15
影响因子：
3.2
通讯作者：
Lund PA
中科院分区：
文献类型：
--
作者：
Sen H;Aggarwal N;Ishionwu C;Hussain N;Parmar C;Jamshad M;Bavro VN;Lund PA
大肠杆菌的EvgS/EvgA双组分系统在低pH和碱金属环境下被激活，并调节许多基因，包括谷氨酸依赖的抗酸系统和一些外排泵的基因。EvgS是大肠杆菌中五种非常规组氨酸激酶（HKs）之一，除了磷酸化受体、HK和二聚化结构域、内部受体和磷酸化转移结构域外，还具有大的周质结构域和细胞质PAS结构域。在pH值为7时组成性激活蛋白的突变映射到PAS结构域。本研究基于BvgS同源物等效区域的结构，构建了EvgS的外质区同源性模型，以指导该区域潜在关键残基的诱变。我们发现组氨酸226是诱导所必需的，并且它在结构上与预测的跨膜螺旋顶部的脯氨酸残基（P522）相匹配，预计在将信息传递到细胞质结构域方面发挥关键作用。我们还表明，PAS结构域的组成突变可以在低外部ph下进一步激活。单独表达该蛋白的细胞质部分也会产生组成激活，如果存在组成PAS突变，则会失去组成激活。这些发现与EvgS感知外部和内部pH值的模型一致，并通过从紧密非活性二聚体到弱活性二聚体的转变而激活，我们提出了对纯化的EvgS细胞质部分的分析，支持这一观点。细菌感知环境的一种方式是通过双组分系统，其中一种膜结合蛋白进行感知，另一种细胞内的膜结合蛋白根据检测到的情况打开或关闭基因。因此，膜结合蛋白必须能够检测到压力，并将这一检测事件发送给细胞内的蛋白质。为了了解这一过程，我们研究了一种蛋白质，这种蛋白质可以帮助大肠杆菌在低pH值环境中存活下来，这是大肠杆菌在胃肠道中定居之前必须做到的。我们描述了蛋白质主要传感部分的预测结构，并确定了其中涉及传感和信号传导过程的一些关键残基。我们提出了一种蛋白质如何被激活的机制，并提出了一些证据来支持我们的建议。
The EvgS/EvgA two-component system of Escherichia coli is activated in response to low pH and alkali metals and regulates many genes, including those for the glutamate-dependent acid resistance system and a number of efflux pumps. EvgS, the sensor kinase, is one of five unconventional histidine kinases (HKs) in E. coli and has a large periplasmic domain and a cytoplasmic PAS domain in addition to phospho-acceptor, HK and dimerization, internal receiver, and phosphotransfer domains. Mutations that constitutively activate the protein at pH 7 map to the PAS domain. Here, we built a homology model of the periplasmic region of EvgS, based on the structure of the equivalent region of the BvgS homologue, to guide mutagenesis of potential key residues in this region. We show that histidine 226 is required for induction and that it is structurally colocated with a proline residue (P522) at the top of the predicted transmembrane helix that is expected to play a key role in passing information to the cytoplasmic domains. We also show that the constitutive mutations in the PAS domain can be further activated by low external pH. Expression of the cytoplasmic part of the protein alone also gives constitutive activation, which is lost if the constitutive PAS mutations are present. These findings are consistent with a model in which EvgS senses both external and internal pH and is activated by a shift from a tight inactive to a weak active dimer, and we present an analysis of the purified cytoplasmic portion of EvgS that supports this. IMPORTANCE One of the ways bacteria sense their environment is through two-component systems, which have one membrane-bound protein to do the sensing and another inside the cell to turn genes on or off in response to what the membrane-bound protein has detected. The membrane-bound protein must thus be able to detect the stress and signal this detection event to the protein inside the cell. To understand this process, we studied a protein that helps E. coli to survive exposure to low pH, which it must do before taking up residence in the gastrointestinal tract. We describe a predicted structure for the main sensing part of the protein and identify some key residues within it that are involved in the sensing and signaling processes. We propose a mechanism for how the protein may become activated and present some evidence to support our proposal.