Analysis of binding domain and function of chimeric mu/kappa opioid receptors to ohmefentanyl stereoisomers

AIM: To investigate specific domains in mu opioid receptors that accounted for selective binding of three stereoisomers of ohmefentanyl (Ohm9204, Ohm9 202, and Ohm9203) and study the function of chimera tt. METHODS: Rat mu and kappa opioid receptors (RMOR, RKOR) and four mu/kappa chimeric receptors ( chimeras) I, II, III, and IV were transiently expressed in COS-1 cells. The binding ability and binding domain of receptor to ligands were determined by radioactive ligand and receptor binding experiments. Through measuring c ellular cAMP levels, we studied the function of chimera If in mediating sig nal transduction. RESULTS: Binding affinities of four chimeric receptors we re similar to wild type opioid receptors RMOR and RKOR). The binding affini ties of Ohm9204 and Ohm9202 to chimera II were similar to that of RMOR. The binding affinities of Ohm9203 to all six receptors were low. U50488 posses sed high binding affinity to chimera I, however dynorphie A(1-9) had some b inding affinity to chimera II that was similar to RKOR, which indicated the domains of RKOR accounting for selectively binding to peptide ligand dynor phie A(1-9) and nonpeptide ligand U50488 were different. The efficacy of Oh m9204 and Ohm9203 on inhibiting forskolin-stimulated cAMP accumulation in c ells transfected with chimera II was similar to that in cells transfected w ith RMOR. CONCLUSION: Replacing 194 - 268 residues of RMOR with 185 - 262 r esidues of RKOR does not influence the ability of mu opioid receptor to bin d Ohm9204 and Ohm9202 and the receptor mediated inhibition of cellular cAMP level.