GPCR engineering yields high-resolution structural insights into β2-adrenergic receptor function

The beta(2)-adrenergic receptor (beta(2)AR) is a well-studied prototype for heterotrimeric guanine nucleotide-binding protein ( G protein)-coupled receptors (GPCRs) that respond to diffusible hormones and neurotransmitters. To overcome the structural flexibility of the b2AR and to facilitate its crystallization, we engineered a b2AR fusion protein in which T4 lysozyme (T4L) replaces most of the third intracellular loop of the GPCR (beta(2)AR-T4L) and showed that this protein retains near-native pharmacologic properties. Analysis of adrenergic receptor ligand-binding mutants within the context of the reported high-resolution structure of beta(2)AR-T4L provides insights into inverse-agonist binding and the structural changes required to accommodate catecholamine agonists. Amino acids known to regulate receptor function are linked through packing interactions and a network of hydrogen bonds, suggesting a conformational pathway from the ligand-binding pocket to regions that interact with G proteins.