Ligand recognition of serine-cysteine amino acid exchanges in transmembrane domain 5 of alpha(2)-adrenergic receptors by UK 14,304

Ligand binding of UK 14,304 reveals notable species (i.e., human-rodent) an d receptor-subtype differences of alpha(2)-adrenergic receptors (alpha(2)-A Rs). To study the molecular basis of the selectivity of UK 14,304, we compa red a series of conservative serine-cysteine exchange mutants at ligand-acc essible positions in transmembrane domain 5 of the human and mouse alpha(2A )-ARs. UK 14,304 bound with similar to 200-fold higher affinity to the huma n alpha(2A)-AR wild-type receptor compared with the human alpha(2A)-ARSer(2 01) mutant, but only an approximately fivefold difference was seen with the corresponding mouse alpha(2A)-AR variant. These effects of cysteine-serine exchanges only involved the agonist low-affinity forms of the receptors, a s the affinity of [H-3]UK 14,304 for the agonist high-affinity receptor pop ulations was not influenced. The apparent affinities of a set of eight stru cturally diverse alpha(2)-AR ligands (six agonists and two antagonists) wer e not influenced significantly by the cysteine-serine exchanges (except for oxymetazoline and yohimbine, with up to nine- and eightfold differences in affinity, respectively). We conclude that position 201 (a) plays a primary role in determining observed subtype/species selectivity of UK 14,304 in c ompetitive antagonist radio-ligand binding assays and (b) does not determin e the subtype selectivity of chlorpromazine.