Galpha(s) and extra-large Galpha(s) (XLalpha(s)) can both transduce receptor activation into intracellular cAMP generation. It is unknown, however, whether these two GNAS-locus products display distinct properties with respect to receptor coupling. Here, we show that XLalpha(s) couples to the beta2-adrenoceptor more efficiently than Galpha(s). In transfected human embryonic kidney 293 cells and mouse embryonic fibroblasts null for both Galpha(s) and XLalpha(s) (2B2 cells), basal cAMP accumulation mediated by XLalpha(s) was higher than that mediated by Galpha(s). Inverse agonist treatment reduced Galpha(s)-mediated basal activity, whereas its effect was markedly blunted on XLalpha(s)-mediated basal activity. Rank order of ligand efficacies regarding cAMP accumulation was the same when the receptor was coupled to XLalpha(s) or Galpha(s). However, ligand-induced and XLalpha(s)-mediated cAMP generation was higher than that mediated by Galpha(s). The relatively high efficiency of XLalpha(s)-mediated cAMP generation was conditional, disappearing with increased level of receptor expression or increased efficacy of ligand. Full-agonist responses in XLalpha(s)- and Galpha(s)-expressing cells were comparable even at low receptor levels, whereas partial agonist responses became comparable only when the receptor expression was increased (>3 pmol/mg). Radioligand binding studies showed that the high-affinity component in agonist binding to beta2-adrenoceptor was more pronounced in cells expressing XLalpha(s) than those expressing Galpha(s). We discuss these findings in the framework of current receptor-G protein activation models and offer an extended ternary complex model that can fully explain our observations.