Characterisation of melanocortin receptor subtypes by radioligand binding analysis

Abstract

The DNAs encoding three melanocortin receptor subtypes (melanocortin MC 1 receptor, melanocortin MC 3 receptor and melanocortin MC 5 receptor) were expressed individually in COS ( C V-1 O rigin, SV40) cells to characterise their ligand binding properties. The results indicated that [ 125I][Nle 4, D-Phe 7]α-MSH (melanocyte stimulating hormone) bound to a single saturable site with K d values of 85.1 ± 8.0 pmol/l (mean ± S.E.M.), 396 ± 65 pmol/l and 5.05 ± 1.00 nmol/l for melanocortin MC 1 receptor, melanocortin MC 3 receptor and melanocortin MC 3 receptor, respectively. The melanocortin MC 1 receptor and the melanocortin MC 5 receptor showed a similar potency order to the melanocortic peptides examined which was markedly different from the potency order of the melanocortin MC 3 receptor. The melanocortin MC 1 receptor and melanocortin MC 5 receptor had a relatively higher affinity for α-MSH than γ-MSH and β-MSH, whereas the melanocortin MC 3 receptor had higher affinity for desacetyl-α-MSH, γ-MSH and β-MSH compared to α-MSH. The inclusion of the endopeptidase inhibitor phosphoramidon to prevent the breakdown of ACTH-(1-39) (adrenocorticotrophic hormone) to α-MSH, decreased ACTH-(1-39) binding affinity showing that ACTH-(1-39) had a much lower affinity for melanocortin MC 1 receptor than reported earlier.