PT - JOURNAL ARTICLE AU - Michael Ippolito AU - Francesco De Pascali AU - Asuka Inoue AU - Jeffrey L. Benovic TI - Phenylalanine 193 in Extracellular Loop 2 of the <em>β</em><sub>2</sub>-Adrenergic Receptor Coordinates <em>β</em>-Arrestin Interaction AID - 10.1124/molpharm.121.000332 DP - 2022 Feb 01 TA - Molecular Pharmacology PG - 87--94 VI - 101 IP - 2 4099 - http://molpharm.aspetjournals.org/content/101/2/87.short 4100 - http://molpharm.aspetjournals.org/content/101/2/87.full SO - Mol Pharmacol2022 Feb 01; 101 AB - 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 β2-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 β2AR-F193A. This single point mutation in extracellular loop 2 of the β2AR is sufficient to intrinsically bias the β2AR away from β-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.SIGNIFICANCE STATEMENT The role of extracellular G protein-coupled receptor (GPCR) domains in mediating intracellular interactions is poorly understood. We characterized the effects of extracellular loop mutations on agonist-promoted interactions of GPCRs with G protein and β-arrestin. Our studies reveal that F193 in extracellular loop 2 in the β2-adrenergic receptor mediates interactions with G protein and β-arrestin with a biased loss of β-arrestin binding. These results provide new insights on the role of the extracellular domain in differentially modulating intracellular interactions with GPCRs.