Department of Physiology & Biophysics, Weill Cornell Medicine, 1300 York Avenue, New York, New York 10021, United States.
Howard Hughes Medical Institute, Chevy Chase, Maryland 20815, United States.
J Am Chem Soc. 2023 Apr 19;145(15):8583-8592. doi: 10.1021/jacs.3c01003. Epub 2023 Apr 6.
Limited chemical shift dispersion represents a significant barrier to studying multistate equilibria of large membrane proteins by F NMR. We describe a novel monofluoroethyl F probe that dramatically increases the chemical shift dispersion. The improved conformational sensitivity and line shape enable the detection of previously unresolved states in one-dimensional (1D) F NMR spectra of a 134 kDa membrane transporter. Changes in the populations of these states in response to ligand binding, mutations, and temperature correlate with population changes of distinct conformations in structural ensembles determined by single-particle cryo-electron microscopy (cryo-EM). Thus, F NMR can guide sample preparation to discover and visualize novel conformational states and facilitate image analysis and three-dimensional (3D) classification.
有限的化学位移分散是研究大膜蛋白的多态平衡的一个重要障碍通过 F NMR。我们描述了一种新的单氟乙基 F 探针，它大大增加了化学位移分散。改进的构象敏感性和线形使以前在一维（1D）中无法分辨的状态得以检测 F NMR 光谱的 134 kDa 膜转运蛋白。这些状态的群体变化对配体结合、突变和温度的响应与通过单颗粒低温电子显微镜（cryo-EM）确定的结构集合中的不同构象的群体变化相关。因此，F NMR 可以指导样品制备，以发现和可视化新的构象状态，并促进图像分析和三维（3D）分类。
