Chemical Research Laboratory, University of Oxford, Oxford, UK.
OMass Therapeutics, Oxford, UK.
Nat Chem. 2022 Dec;14(12):1375-1382. doi: 10.1038/s41557-022-01041-9. Epub 2022 Nov 10.
G-protein-coupled receptors signal through cognate G proteins. Despite the widespread importance of these receptors, their regulatory mechanisms for G-protein selectivity are not fully understood. Here we present a native mass spectrometry-based approach to interrogate both biased signalling and allosteric modulation of the β-adrenergic receptor in response to various ligands. By simultaneously capturing the effects of ligand binding and receptor coupling to different G proteins, we probed the relative importance of specific interactions with the receptor through systematic changes in 14 ligands, including isoprenaline derivatives, full and partial agonists, and antagonists. We observed enhanced dynamics of the intracellular loop 3 in the presence of isoprenaline, which is capable of acting as a biased agonist. We also show here that endogenous zinc ions augment the binding in receptor-G complexes and propose a zinc ion-binding hotspot at the TM5/TM6 intracellular interface of the receptor-G complex. Further interrogation led us to propose a mechanism in which zinc ions facilitate a structural transition of the intermediate complex towards the stable state.
G 蛋白偶联受体通过同源 G 蛋白传递信号。尽管这些受体具有广泛的重要性，但它们对 G 蛋白选择性的调节机制尚未完全了解。在这里，我们提出了一种基于天然质谱的方法，用于检测β-肾上腺素能受体在不同配体作用下的偏向信号和变构调节。通过同时捕捉配体结合和受体与不同 G 蛋白偶联的影响，我们通过系统改变 14 种配体（包括异丙肾上腺素衍生物、完全激动剂、部分激动剂和拮抗剂）来探测与受体特异性相互作用的相对重要性。我们观察到异丙肾上腺素存在时细胞内环 3 的动态增强，异丙肾上腺素能够作为一种偏向激动剂发挥作用。我们还在此表明，内源性锌离子增强了受体-G 复合物中的结合，并提出了受体-G 复合物 TM5/TM6 细胞内界面处的锌离子结合热点。进一步的研究使我们提出了一种机制，其中锌离子促进中间复合物向稳定状态的结构转变。
