A Selective Tether Recruits Activated Response Regulator CheB to Its Chemoreceptor Substrate.
A Selective Tether Recruits Activated Response Regulator CheB to Its Chemoreceptor Substrate.
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DOI：
10.1128/mbio.03106-21
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发表时间：
2021-12-21
期刊：
影响因子：
6.4
通讯作者：
Hazelbauer GL
中科院分区：
文献类型：
--
作者：
Li M;Xu X;Zou X;Hazelbauer GL
甲基酯酶/脱酰胺酶CheB是细菌趋化系统的关键组成部分。它也是双组分响应调节器的一个突出例子，其中效应域是一种酶。与其他应答调节因子一样，CheB通过其调节结构域的天冬氨酸残基磷酸化激活，在其两个结构域之间形成开放构象。在大肠杆菌和相关生物中对CheB的研究表明，在化学受体羧基末端的酶的五肽结合位点也增强了CheB的酶促作用。相关的羧基末端五肽在分布于11个细菌门和许多细菌种的bb50 000个化学受体序列中被发现，它们可能在其中起着相似的作用。然而，关于CheB磷酸化、五肽结合及其与底物甲基酯和酰胺在化学受体体上的相互作用的相互关系知之甚少。我们通过表征CheB与纳米盘插入的化学受体二聚体的结合动力学进行了研究。由此得到的动力学和热力学常数揭示了CheB磷酸化与五肽结合之间的协同作用，其中磷酸化模拟了五肽的结合，五肽不仅作为CheB的高亲和力系链，而且还选择了酶的激活构象。这种选择的基础是通过分子模型揭示，预测CheB上的五肽结合位点仅存在于开放的活化酶中。通过选择性捆绑来招募活化酶代表了一种以前未被重视的调节反应调节作用的策略，这种策略很可能发生在其他双组分系统中。
Methylesterase/deamidase CheB is a key component of bacterial chemotaxis systems. It is also a prominent example of a two-component response regulator in which the effector domain is an enzyme. Like other response regulators, CheB is activated by phosphorylation of an aspartyl residue in its regulatory domain, creating an open conformation between its two domains. Studies of CheB in Escherichia coli and related organisms have shown that its enzymatic action is also enhanced by a pentapeptide-binding site for the enzyme at the chemoreceptor carboxyl terminus. Related carboxyl-terminal pentapeptides are found on >25,000 chemoreceptor sequences distributed across 11 bacterial phyla and many bacterial species, in which they presumably play similar roles. Yet, little is known about the interrelationship of CheB phosphorylation, pentapeptide binding, and interactions with its substrate methylesters and amides on the body of the chemoreceptor. We investigated by characterizing the binding kinetics of CheB to Nanodisc-inserted chemoreceptor dimers. The resulting kinetic and thermodynamic constants revealed a synergy between CheB phosphorylation and pentapeptide binding in which a phosphorylation mimic enhanced pentapeptide binding, and the pentapeptide served not only as a high-affinity tether for CheB but also selected the activated conformation of the enzyme. The basis of this selection was revealed by molecular modeling that predicted a pentapeptide-binding site on CheB which existed only in the open, activated enzyme. Recruitment of activated enzyme by selective tethering represents a previously unappreciated strategy for regulating response regulator action, one that may well occur in other two-component systems.