Phenylalanine 193 in Extracellular Loop 2 of the β2-Adrenergic Receptor Coordinates β-Arrestin Interaction

G protein-coupled receptors (GPCRs) transduce a diverse variety of extracellular stimuli into intracellular signaling. These receptors are the most clinically productive drug targets at present. Despite decades of research on the signaling consequences of molecule-receptor interactions, conformational components of receptor-effector interactions remain incompletely described. The b2-adrenergic receptor (β2AR) is a prototypical and extensively studied GPCR that can provide insight into this aspect of GPCR signaling thanks to robust structural data and rich pharmacopeia. Using bioluminescence resonance energy transfer -based biosensors, second messenger assays, and biochemical techniques, we characterize the properties of b2AR-F193A. This single point mutation in extracellular loop 2 of the b2AR is sufficient to intrinsically bias the b2AR away from b-arrestin interaction and demonstrates altered regulatory outcomes downstream of this functional selectivity. This study highlights the importance of extracellular control of intracellular response to stimuli and suggests a previously undescribed role for the extracellular loops of the receptor and the extracellular pocket formed by transmembrane domains 2, 3, and 7 in GPCR regulation that may contribute to biased signaling at GPCRs.