Phosphorylation of the -Opioid Receptor at Tyrosine 166 (Tyr3.51) in the DRY Motif Reduces Agonist Efficacy□S

The effects of phosphorylation of the tyrosine residue in the highly conserved DRY motif expressed in the putative second cytoplasmic loop of the -opioid receptor were assessed after expression in human embryonic kidney (HEK) 293 cells. Ty-rosine kinase activation by epidermal growth factor (EGF) or hydrogen peroxide treatment effectively increased phosphory-lation of the tyrosine-166 in the -opioid receptor (MOR-Tyr166p) as measured by a novel phosphoselective antibody. We were surprised to find that the increase in MOR-Tyr166p immunoreactivity (ir) required coactivation by the opioid agonist [D-Ala2,methyl-Phe4,Gly5-ol]enkephalin (DAMGO), as demon-strated by both Western blot imaging of membrane proteins and confocal microscopy of transfected cells; MOR-Tyr166p-ir did not significantly increase after either DAMGO, EGF, or H2O