The human delta opioid receptor (hdOR) was used as a model to study the role of ER calcium pump, Sarco(endo)plasmic reticulum ATPase 2b (SERCA2b), in the biogenesis of G protein-coupled receptors (GPCRs). Stable cell lines with inducible expression of the hdOR and wild-type or mutant SERCA2b with various tags were created. Transient transfections were used for co-expression, interactions were examined by co-immunoprecipitation of native or chemically crosslinked proteins, receptor maturation was followed by pulse-chase assay and Fluo-3/AM was used to measure ER-released calcium by confocal microscopy. We have earlier reported that newly synthesized hdOR precursors interact with SERCA2b and also with ER molecular chaperone calnexin. Calnexin has also been suggested to modulate SERCA2b activity in vivo. Here we show that SERCA2b, calnexin and the receptor precursor form a ternary complex that is not dependent on receptor N-glycans. Different domains of SERCA2b mediate the interaction with calnexin and the receptor. Also, calcium and ATP had a different modulatory effect on the interactions. The co-expression of catalytically inactive SERCA2b mutant (D351A) reduced receptor maturation without a collapse in ER calcium level. The decrease in hdOR maturation was similar to the effect of SERCA inhibitor thapsigargin (Tg). However, unlike Tg, mutation of the active site of SERCA2b, which stabilizes a different conformation of the calcium pump, did not abolish receptor interaction. We conclude that the active SERCA2b modulates GPCR biogenesis via dynamic protein-protein interactions.