Conformational coupling between receptor and kinase binding sites through a conserved salt bridge in a signaling complex scaffold protein.
Conformational coupling between receptor and kinase binding sites through a conserved salt bridge in a signaling complex scaffold protein.
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DOI：
10.1371/journal.pcbi.1003337
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发表时间：
2013
影响因子：
4.3
通讯作者：
Zhulin IB
中科院分区：
文献类型：
--
作者：
Ortega DR;Mo G;Lee K;Zhou H;Baudry J;Dahlquist FW;Zhulin IB
细菌的趋化性是目前研究最多的信号转导途径之一。CheW是介导化学感受器和CheA激酶在三元信号传导复合物中缔合的支架蛋白。用其他残基取代CheW的保守Arg62的效果表明，支架蛋白发挥着比简单地结合其伴侣蛋白更复杂的作用。虽然R62A CheW对化学感受器和CheA具有基本相同的亲和力，但表达突变蛋白的细胞在趋化性方面受损。使用分子动力学模拟（MD），NMR光谱和圆二色性（CD）的组合，我们解决了Arg62的作用。在这里，我们表明，Arg62形成一个盐桥与另一个高度保守的残基，Glu38。虽然这种相互作用对整体蛋白质稳定性并不重要，但保持CheW的化学感受器和激酶结合位点的正确对齐至关重要。计算和实验数据表明，盐桥在维持两个伴侣结合位点的对齐中的作用是信号复合物的功能的基础，但不是其组装。我们得出结论，咀嚼的一个关键特征是保持特定的几何形状之间的两个相互作用的网站所需的功能作为一个支架。信号转导是一个普遍的生物学过程，也是药物设计的共同目标。大肠杆菌的趋化机制是一个典型的信号转导系统，而CheW蛋白是其核心成分之一。CheW被认为是一种支架蛋白，介导与CheA组氨酸激酶和化学受体形成信号复合物。针对高度保守的残基Arg62的突变损害趋化性，同时保持对两个伙伴的正常结合亲和力，这表明Chew可能发挥比以前提出的更复杂的作用。利用一系列的分子动力学模拟，我们发现残基Arg62可以与另一个高度保守的残基Glu38形成稳定的盐桥。我们确定该桥不有助于蛋白质的整体稳定性。然而，桥稳定的本地骨干结构的咀嚼和稳定的化学受体和激酶的结合位点的相对位置。这些相互作用的几何形状似乎对信号复合物的功能至关重要。我们使用NMR光谱和圆二色性分析验证并补充了我们的计算结果。
Bacterial chemotaxis is one of the best studied signal transduction pathways. CheW is a scaffold protein that mediates the association of the chemoreceptors and the CheA kinase in a ternary signaling complex. The effects of replacing conserved Arg62 of CheW with other residues suggested that the scaffold protein plays a more complex role than simply binding its partner proteins. Although R62A CheW had essentially the same affinity for chemoreceptors and CheA, cells expressing the mutant protein are impaired in chemotaxis. Using a combination of molecular dynamics simulations (MD), NMR spectroscopy, and circular dichroism (CD), we addressed the role of Arg62. Here we show that Arg62 forms a salt bridge with another highly conserved residue, Glu38. Although this interaction is unimportant for overall protein stability, it is essential to maintain the correct alignment of the chemoreceptor and kinase binding sites of CheW. Computational and experimental data suggest that the role of the salt bridge in maintaining the alignment of the two partner binding sites is fundamental to the function of the signaling complex but not to its assembly. We conclude that a key feature of CheW is to maintain the specific geometry between the two interaction sites required for its function as a scaffold. Signal transduction is a universal biological process and a common target of drug design. The chemotaxis machinery in Escherichia coli is a model signal transduction system, and the CheW protein is one of its core components. CheW is thought to work as a scaffold protein that mediates the formation of the signaling complex with the CheA histidine kinase and the chemoreceptors. A mutation targeting a highly conserved residue, Arg62, impairs chemotaxis while maintaining normal binding affinity for both partners, suggesting that CheW might play a more complex role than previously proposed. Using a series of molecular dynamics simulations, we found that the residue Arg62 can form a stable salt bridge with another highly conserved residue, Glu38. We determined that this bridge does not contribute to the overall stability of the protein. However, the bridge stabilizes the local backbone structure of CheW and stabilizes the relative position of the binding sites for the chemoreceptor and kinase. The geometry of these interactions appears to be vital for the function of the signaling complex. We validated and complemented our computational findings using NMR spectroscopy and circular dichroism analysis.
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影响因子：
3.6
作者：
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通讯作者：
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