The role of intrinsically disordered domains in regulating G protein coupled receptor signaling

The α _2A adrenergic receptor (α _2A AR) is a clinically important target for various diseases including hypertension, diabetes and chronic pain. Here, using single-molecule fluorescence resonance energy transfer imaging, we show how agonist-specific activation dynamics in both structured transmembrane domain (TMD) and intrinsically disorders regions (IDRs) of α _2A AR lead to diverse signaling profiles. Through seven pairs of strategically designed fluorophore labels, we systematically investigate the real-time conformational changes of α _2A AR. Our study reveals unique TM6 dynamics in α _2A AR, featured by a high energy barrier for agonist-induced outward movements essential for activation. In contrast, we identify agonist-specific conformational dynamics of a partially disordered extracellular loop (ECL2), highlighting its role as a dynamic regulatory module that controls receptor function. Moreover, we characterize the conformational landscapes of the long third intracellular loop (ICL3), revealing its compact structural features and membrane-proximal localization in the basal state, where it acts as a negative allosteric regulator in transducer coupling. Furthermore, we identify multiful functional sub-states of ICL3 that are dynamically modulated by both kinase phosphorylation and drug efficacy. These findings offer previously underappreciated structural and dynamic insights into α _2A AR function governed by both TMD and IDRs, and may open up new avenues for the development of better therapeutics.