MAPPING OF DOPAMINE D-3 RECEPTOR-BINDING SITE BY PHARMACOLOGICAL CHARACTERIZATION OF MUTANTS EXPRESSED IN CHO CELLS WITH THE SEMLIKI-FOREST-VIRUS SYSTEM

Nine mutants and the wild-type human dopamine D-3 receptor were expres sed at high levels in BHK and CHO cells using the Semliki Forest virus system and were analysed for receptor binding with several structural ly different dopamine D-3 ligands. The mutation His349Leu showed a sig nificant decrease in pK(i) values for raclopride, dopamine and GR21823 1, but an increase in affinity for GR99841. Thr369Val had an increase in pK(i) for both GR99841 and 7-OH-DPAT. The receptor modelling based on sequence alignment with bacteriorhodopsin indicated that Thr369 and His349 are located on the inside of the ligand binding pocket and the effect of the mutagenesis was therefore expected. The change in bindi ng affinity for Thr369Val could be due to the location in the transmem brane domain VII close to the aspartate residue in domain III, the pos tulated counter ion for dopamine.