Abstract

Chimeric receptors of the β₂-adrenergic receptor in which the extracellular loops were replaced with the corresponding amino acids of the α_1a-adrenergic receptor were generated to measure changes in α₁-antagonist affinity. Although no changes in α₁-antagonist affinity were measured in the β₂/α_1a chimeras, a decreased IC₅₀ (10-fold) for agonists as compared with wild type β₂ control was found because of the replacement of the third extracellular loop (EX3). These agonist high affinity changes were because of a greater proportion of high affinity sites (2-fold) that were convertible to low affinity sites with guanosine 5′-3-O-(thio)triphosphate. Adenylate cyclase activity evoked by the EX3 chimera showed commensurate increases in the basal signal transduction as well as the isoproterenol-stimulated potency, suggesting constitutive activity. However, unlike other constitutively active adrenergic receptor mutants in which the mutation causes G protein-independent changes, the mechanism of the EX3 chimera seems to be attributable to a greater ease with which the active ternary complex is formed because of a higher affinity/coupling of the G protein. Although the changes because of EX3 are indirect and most likely affect helical packing, they support an emerging hypothesis that G protein-coupled receptors have evolved their structure-function relationships to constrain the receptor in an inactive state.