Non‐canonical activation of histidine kinase KdpD by phosphotransferase protein PtsN through interaction with the transmitter domain
Non‐canonical activation of histidine kinase KdpD by phosphotransferase protein PtsN through interaction with the transmitter domain
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DOI：
10.1111/mmi.13751
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发表时间：
2017-10
影响因子：
3.6
通讯作者：
Markus Mörk-Mörkenstein;R. Heermann;Yvonne Göpel;K. Jung;B. Görke
中科院分区：
文献类型：
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作者：
Markus Mörk-Mörkenstein;R. Heermann;Yvonne Göpel;K. Jung;B. Görke
KdpD/KdpE双组分系统通过控制高亲和力K+转运体KdpFABC的合成来调控K+的动态平衡。当感受到环境中的低K+浓度时，二聚体蛋白KdpD自动磷酸化反式，并将磷酸化基团转移到反应调节因子KdpE，KdpE随后激活kdpFABC转录。在大肠杆菌中，KdpD也可以通过与辅助蛋白PtsN的非磷酸化形式相互作用而被激活。PtsN刺激KdpD激酶活性，从而增加磷酸化KdpE水平。在这里，我们分析了KdpD/KdpE和PtsN之间的相互作用。PtsN与KdpD的催化DHP结构域特异结合，KdpE也与该结构域接触。因此，PtsN和KdpE竞争绑定，形成了一个悖论。低水平的非磷酸化PtsN刺激KdpFABC的表达，而高水平的PtsN则通过阻断KdpE对KdpD的通路而降低kdpFABC的表达。配体钓鱼实验提供了洞察力，因为他们揭示了PtsN/KdpD2/KdpE在体内形成的三元络合物，表明PtsN和KdpE结合了KdpD二聚体中的不同原型。PtsN可以结合一个原基来刺激第二个KdpD原基的磷酸化，然后第二个KdpD原基磷酸化结合KdpE。PtsN的磷酸化阻止了它在三元络合物中的掺入。与保守的DHP结构域的相互作用使PtsN能够调节额外的激酶，如phor。
The two‐component system KdpD/KdpE governs K+ homeostasis by controlling synthesis of the high affinity K+ transporter KdpFABC. When sensing low environmental K+ concentrations, the dimeric kinase KdpD autophosphorylates in trans and transfers the phosphoryl‐group to the response regulator KdpE, which subsequently activates kdpFABC transcription. In Escherichia coli, KdpD can also be activated by interaction with the non‐phosphorylated form of the accessory protein PtsN. PtsN stimulates KdpD kinase activity thereby increasing phospho‐KdpE levels. Here, we analyzed the interplay between KdpD/KdpE and PtsN. PtsN binds specifically to the catalytic DHp domain of KdpD, which is also contacted by KdpE. Accordingly, PtsN and KdpE compete for binding, providing a paradox. Low levels of non‐phosphorylated PtsN stimulate, whereas high amounts reduce kdpFABC expression by blocking access of KdpE to KdpD. Ligand fishing experiments provided insight as they revealed ternary complex formation of PtsN/KdpD2/KdpE in vivo demonstrating that PtsN and KdpE bind different protomers in the KdpD dimer. PtsN may bind one protomer to stimulate phosphorylation of the second KdpD protomer, which then phosphorylates bound KdpE. Phosphorylation of PtsN prevents its incorporation in ternary complexes. Interaction with the conserved DHp domain enables PtsN to regulate additional kinases such as PhoR.