National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD, USA.
Methods Mol Biol. 2021;2268:61-76. doi: 10.1007/978-1-0716-1221-7_4.
G protein-coupled receptors (GPCR) are integral membrane proteins that regulate multiple cellular processes. To obtain insights into structural properties of GPCR and mechanism of activity, these proteins should be isolated in significant (milligram) quantities, in a pure, homogenous, and stable form. Here we describe the expression and purification of type II human cannabinoid receptor CB, a class A GPCR, in two different types of expression hosts: in Escherichia coli and in mammalian suspension cell culture Expi293. Our method allows preparation of milligram quantities of the purified receptors suitable for a wide array of downstream applications including high-resolution structural studies and functional assays.
G 蛋白偶联受体（GPCR）是调节多种细胞过程的完整膜蛋白。为了深入了解 GPCR 的结构特性和活性机制，这些蛋白质应以大量（毫克级）、纯、均一和稳定的形式分离出来。在这里，我们描述了在两种不同类型的表达宿主（大肠杆菌和哺乳动物悬浮细胞培养物 Expi293）中表达和纯化 II 型人大麻素受体 CB（一种 A 类 GPCR）的方法。我们的方法可以制备毫克级的纯化受体，适用于广泛的下游应用，包括高分辨率结构研究和功能测定。
