title: Co-operative regulation of ligand binding to melanocortin receptor subtypes: evidence for interacting binding sites.
abstract: This study evaluates the binding the melanocyte stimulating hormone peptide analogue [125I]NDP-MSH to melanocortin receptors MC1, MC3, MC4 and MC5 in insect cell membranes produced by baculovirus expression systems. The presence of Ca2+ was found to be mandatory to achieve specific [125I]NDP-MSH binding to the melanocortin receptors. Although association kinetics of [125I]NDP-MSH followed the regularities of simple bimolecular reactions, the dissociation of [125I]NDP-MSH from the melanocortin receptors was heterogeneous. Eleven linear and cyclic MSH peptides studied displaced the [125I]NDP-MSH binding to the studied melanocortin receptors, with the shapes of their competition curves varying from biphasic or shallow to super-steep (Hill coefficients ranging from 0.4 to 1.5). Notably the same peptide often gave highly different patterns on different melanocortin receptor subtypes; e.g. the MC4 receptor selective antagonist HS131 gave a Hill coefficient of 1.5 on the MC1 receptor but 0.5-0.7 on the MC(3-5) receptors. Adding a mask of one of the peptides to block its high affinity binding did not prevent other competing peptides to yield biphasic competition curves. The data indicate that the binding of MSH peptides to melanocortin receptors are governed by a complex dynamic homotropic co-operative regulations.
authors.affiliation: Department of Pharmaceutical Biosciences, Uppsala University, BMC Box 591, 751 24 Uppsala, Sweden.
authors.forename: Sergei
authors.initials: S
authors.lastname: Kopanchuk
authors.forename: Santa
authors.initials: S
authors.lastname: Veiksina
authors.forename: Ramona
authors.initials: R
authors.lastname: Petrovska
authors.forename: Ilze
authors.initials: I
authors.lastname: Mutule
authors.forename: Michael
authors.initials: M
authors.lastname: Szardenings
authors.forename: Ago
authors.initials: A
authors.lastname: Rinken
authors.forename: Jarl E S
authors.initials: JE
authors.lastname: Wikberg
mesh_terms: D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D007457:Iodine Radioisotopes; D007700:Kinetics; D008024:Ligands; D008546:Melanoma, Experimental / Q000378:metabolism / Q000473:pathology; D010456:Peptides, Cyclic / Q000378:metabolism / Q000494:pharmacology; D011869:Radioligand Assay; D044102:Receptor, Melanocortin, Type 1 / Q000378:metabolism; D044104:Receptor, Melanocortin, Type 3 / Q000378:metabolism; D044105:Receptor, Melanocortin, Type 4 / Q000378:metabolism; D044101:Receptors, Melanocortin / Q000378:metabolism*; D018411:Spodoptera; D013997:Time Factors; D000521:alpha-MSH / Q000031:analogs & derivatives* / Q000378:metabolism; D019825:gamma-MSH / Q000378:metabolism / Q000494:pharmacology
chemical_list: C472863:HS 131; D007457:Iodine Radioisotopes; D008024:Ligands; D010456:Peptides, Cyclic; D044102:Receptor, Melanocortin, Type 1; D044104:Receptor, Melanocortin, Type 3; D044105:Receptor, Melanocortin, Type 4; D044101:Receptors, Melanocortin; D019825:gamma-MSH; D000521:alpha-MSH; C027756:MSH, 4-Nle-7-Phe-alpha-; D002118:Calcium