Biased agonism of carvedilol in the beta1-adrenergic receptor is governed by conformational exclusion

Abstract

Only two non-visual arrestins recognize many hundreds of different, intracellularly phosphorylated G protein-coupled receptors (GPCRs). Due to the highly dynamic nature of GPCR*arrestin complexes, the critical determinants of GPCR-arrestin recognition have remained largely unclear. We show here that arrestin2 recruitment to the {beta}1-adrenergic receptor ({beta}1AR) can be induced by an arrestin-activating phosphopeptide that is not covalently linked to the receptor and that the recruitment is independent of the presence and type of the orthosteric receptor ligand. Apparently, the arrestin-receptor interaction is driven by the conformational switch within arrestin induced by the phosphopeptide, whereas the electrostatic attraction towards the receptor phosphosites may only play an auxiliary role. Extensive NMR observations show that in contrast to previous static GPCR*arrestin complex structures, the {beta}1AR complex with the beta-blocker carvedilol and arrestin2 is in a G protein-inactive conformation. The insensitivity to the specific receptor conformation provides a rationale for arrestins promiscuous recognition of GPCRs and explains the arrestin-biased agonism of carvedilol, which largely blocks G protein binding, while still enabling arrestin engagement. Significance statementG protein-coupled receptors regulate cellular signaling through G proteins and arrestins. While G protein interactions are well understood, the molecular basis of arrestin recognition remains unclear due to the dynamic nature of GPCR*arrestin complexes. We show that arrestin recognition of the {beta}1-adrenergic receptor occurs independently of the receptors conformational state or ligand binding. Using NMR, cryo-EM, and biochemical assays, we find that arrestin engagement is driven by a conformational change within arrestin itself, triggered by a non-receptor-attached phosphopeptide, whereas electrostatic attraction towards receptor phosphosites may only play an auxiliary role. These findings provide new insights into arrestin activation, explain its ability to recognize diverse GPCRs, with significant implications for understanding biased signaling mechanisms and designing of selective therapeutic strategies.