Department of Chemistry, University of Massachusetts Amherst, Amherst, MA 01003, United States.
Department of Chemistry, University of Massachusetts Amherst, Amherst, MA 01003, United States.
Biochim Biophys Acta Biomembr. 2020 Sep 1;1862(9):183312. doi: 10.1016/j.bbamem.2020.183312. Epub 2020 Apr 15.
Bacterial chemoreceptors are organized in arrays composed of helical receptors arranged as trimers of dimers, coupled to a histidine kinase CheA and a coupling protein CheW. Ligand binding to the external domain inhibits the kinase activity, leading to a change in the swimming behavior. Adaptation to an ongoing stimulus involves reversible methylation and demethylation of specific glutamate residues. However, the exact mechanism of signal propagation through the helical receptor to the histidine kinase remains elusive. Dynamics of the receptor cytoplasmic domain is thought to play an important role in the signal transduction, and current models propose inverse dynamic changes in different regions of the receptor. We hypothesize that the adaptational modification (methylation) controls the dynamics by stabilizing a partially ordered domain, which in turn modulates the binding of the kinase, CheA. We investigated the difference in dynamics between the methylated and unmethylated states of the chemoreceptor using solid-state NMR. The unmethylated receptor (CF4E) shows increased flexibility relative to the methylated mimic (CF4Q). Methylation helix 1 (MH1) has been shown to be flexible in the methylated mimic receptor. Our analysis indicates that in addition to MH1, methylation helix 2 also becomes flexible in the unmethylated receptor. In addition, we have demonstrated that both states of the receptor have a rigid region and segments with intermediate timescale dynamics. The strategies used in this study for identifying dynamic regions are applicable to a broad class of proteins and protein complexes with intrinsic disorder and dynamics spanning multiple timescales.
细菌化学感受器以由二聚体三聚体组成的螺旋受体排列的阵列形式存在，与组氨酸激酶 CheA 和偶联蛋白 CheW 相偶联。配体与外域结合抑制激酶活性，导致游泳行为发生变化。对持续刺激的适应涉及特定谷氨酸残基的可逆甲基化和去甲基化。然而，通过螺旋受体将信号传递到组氨酸激酶的确切机制仍不清楚。受体胞质域的动力学被认为在信号转导中起着重要作用，目前的模型提出了受体不同区域的反向动力学变化。我们假设适应性修饰（甲基化）通过稳定部分有序结构域来控制动力学，从而调节激酶 CheA 的结合。我们使用固态 NMR 研究了化学感受器的甲基化和非甲基化状态之间的动力学差异。未甲基化的受体（CF4E）与甲基化模拟物（CF4Q）相比显示出更高的灵活性。已经表明，甲基化螺旋 1（MH1）在甲基化模拟物受体中是灵活的。我们的分析表明，除了 MH1 之外，甲基化螺旋 2 在未甲基化受体中也变得灵活。此外，我们已经证明受体的两种状态都具有刚性区域和具有中间时间尺度动力学的片段。本研究中用于识别动态区域的策略适用于具有固有无序性和跨越多个时间尺度的动力学的广泛的蛋白质和蛋白质复合物。
