Specificity residues determine binding affinity for two-component signal transduction systems.
Specificity residues determine binding affinity for two-component signal transduction systems.
复制标题
特异性残基决定双组分信号转导系统的结合亲和力。
作者：
Willett,JonathanW;Tiwari,Nitija;Müller,Susanne;Hummels,KatherineR;Houtman,JonCD;Fuentes,ErnestoJ;Kirby,JohnR
双组分系统（TCS）包含组氨酸激酶及其同源反应调节剂，并允许细菌对各种信号进行感知和响应。组氨酸激酶（HK）在对刺激的反应中磷酸化和去磷酸化其同源反应调节因子（RR）。一般来说，这些反应似乎是高度特异性的，需要HK和RR蛋白之间的适当关联。黄色粘球菌基因组编码细菌中最大的信号蛋白库之一（685个开放阅读框[ORF]），包括至少127个HK和至少143个RR。其中，27 arebona fideNtrC家族反应调节因子，其中21个编码邻近其预测的同源激酶。使用全系统分析方法，我们确定HK-NtrC RR对在磷酸转移和磷酸酶功能期间显示动力学偏好，从而定义M. xanthus等温滴定量热法测量表明，同源HK-RR对相互作用的解离常数（Kd）约为1 µM，而非同源对没有可测量的结合。最后，组氨酸激酶，CrdS，和HK 1190之间产生的嵌合体显示，残基赋予磷酸转移和磷酸酶特异性决定结合亲和力，从而建立离散的蛋白质-蛋白质相互作用，防止串扰。这些数据表明，结合亲和力是一个关键的参数管理系统范围内的信号保真度为细菌信号transductionprotein.IMPORTANCEUsingin vitrophosphotransfer和磷酸酶分析和等温滴定量热法，我们已经采取了一个系统范围的方法来证明特异性的一个家庭的两个组件的信号蛋白在粘球菌xanthus。我们的研究结果表明，以前确定的特异性残基决定结合亲和力和磷酸酶特异性遵循磷酸转移特异性同源HK-RR对。数据表明，优先结合亲和力是细菌双组分系统中信号保真度的基础。
Two-component systems (TCS) comprise histidine kinases and their cognate response regulators and allow bacteria to sense and respond to a wide variety of signals. Histidine kinases (HKs) phosphorylate and dephosphorylate their cognate response regulators (RRs) in response to stimuli. In general, these reactions appear to be highly specific and require an appropriate association between the HK and RR proteins. The Myxococcus xanthus genome encodes one of the largest repertoires of signaling proteins in bacteria (685 open reading frames [ORFs]), including at least 127 HKs and at least 143 RRs. Of these, 27 arebona fideNtrC-family response regulators, 21 of which are encoded adjacent to their predicted cognate kinases. Using system-wide profiling methods, we determined that the HK-NtrC RR pairs display a kinetic preference during both phosphotransfer and phosphatase functions, thereby defining cognate signaling systems in M. xanthus. Isothermal titration calorimetry measurements indicated that cognate HK-RR pairs interact with dissociation constants (Kd) of approximately 1 µM, while noncognate pairs had no measurable binding. Lastly, a chimera generated between the histidine kinase, CrdS, and HK1190 revealed that residues conferring phosphotransfer and phosphatase specificity dictate binding affinity, thereby establishing discrete protein-protein interactions which prevent cross talk. The data indicate that binding affinity is a critical parameter governing system-wide signaling fidelity for bacterial signal transduction proteins.IMPORTANCEUsingin vitrophosphotransfer and phosphatase profiling assays and isothermal titration calorimetry, we have taken a system-wide approach to demonstrate specificity for a family of two-component signaling proteins in Myxococcus xanthus. Our results demonstrate that previously identified specificity residues dictate binding affinity and that phosphatase specificity follows phosphotransfer specificity for cognate HK-RR pairs. The data indicate that preferential binding affinity is the basis for signaling fidelity in bacterial two-component systems.