iHuman Institute, ShanghaiTech University, Shanghai 201210, China.
School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China.
Molecules. 2023 Jul 14;28(14):5419. doi: 10.3390/molecules28145419.
Eight hundred and twenty-six human G protein-coupled receptors (GPCRs) mediate the actions of two-thirds of the human hormones and neurotransmitters and over one-third of clinically used drugs. Studying the structure and dynamics of human GPCRs in lipid bilayer environments resembling the native cell membrane milieu is of great interest as a basis for understanding structure-function relationships and thus benefits continued drug development. Here, we incorporate the human A adenosine receptor (AAR) into lipid nanodiscs, which represent a detergent-free environment for structural studies using nuclear magnetic resonance (NMR) in solution. The [N,H]-TROSY correlation spectra confirmed that the complex of [u-N, ~70% H]-AAR with an inverse agonist adopts its global fold in lipid nanodiscs in solution at physiological temperature. The global assessment led to two observations of practical interest. First, AAR in nanodiscs can be stored for at least one month at 4 °C in an aqueous solvent. Second, LMNG/CHS micelles are a very close mimic of the environment of AAR in nanodiscs. The NMR signal of five individually assigned tryptophan indole N-H moieties located in different regions of the receptor structure further enabled a detailed assessment of the impact of nanodiscs and LMNG/CHS micelles on the local structure and dynamics of AAR. As expected, the largest effects were observed near the lipid-water interface along the intra- and extracellular surfaces, indicating possible roles of tryptophan side chains in stabilizing GPCRs in lipid bilayer membranes.
八百二十六个人类 G 蛋白偶联受体 (GPCR) 介导了三分之二的人类激素和神经递质以及三分之一以上的临床应用药物的作用。在类似于天然细胞膜环境的脂质双层环境中研究人类 GPCR 的结构和动力学，对于理解结构-功能关系非常重要，因此有利于持续的药物开发。在这里，我们将人类 A 腺苷受体 (AAR) 纳入脂质纳米盘，这是一种在溶液中使用核磁共振 (NMR) 进行结构研究的无去污剂环境。[N,H]-TROSY 相关谱证实，与反向激动剂复合的 [u-N,~70% H]-AAR 采用其在生理温度下溶液中脂质纳米盘中的整体折叠。整体评估得出了两个具有实际意义的观察结果。首先，AAR 在纳米盘中在 4°C 下可在水性溶剂中至少保存一个月。其次，LMNG/CHS 胶束非常接近 AAR 在纳米盘中的环境。五个单独分配的色氨酸吲哚 N-H 部分的 NMR 信号位于受体结构的不同区域，进一步使我们能够详细评估纳米盘和 LMNG/CHS 胶束对 AAR 的局部结构和动力学的影响。如预期的那样，在跨膜脂质双层的内表面和外表面附近观察到最大的影响，表明色氨酸侧链可能在稳定 GPCR 方面发挥作用脂质双层膜。
