Effects of amino acid substitutions in the first extracellular loop region of the delta- and mu-opioid receptors were examined. Substitution of lysine-108 of the delta-receptor (delta K108) with asparagine improved affinity to [D-Ala(2),MePhe(4),Gly-ol(5)]enk ephalin (DAGO), a mu-selective peptide agonist, to be comparable with that of the mu-receptor. On the other hand, replacement of mN127 with lysine decreased the affinity to DAGO by similar to 15-fold. These results suggest that dK108 and mN127, which correspond to each other in the aligned amino acid sequences, mainly determine the difference in DAGO binding affinity between the delta- and mu-receptors.