Serine residues 110 and 114 are required for agonist binding but not antagonist binding to the melatonin MT(1) receptor.
Site-directed mutation of serine 110 (Ser(3.35)) and serine 114 (Ser(3.39)) in the human melatonin MT(1) receptor to alanine residues reduced ligand binding affinities of seven known melatonin receptor agonists and partial agonists by 3- to 15-fold. These mutants also displayed a relative reduction in their affinities for melatonin-mediated functional responses of 30- and 14-fold, respectively. In contrast to the observed effects of the agonists and partial agonists, the melatonin receptor antagonist luzindole was found to bind to mutants Ser(3.35)Ala and Ser(3.39)Ala with affinities equivalent to that determined for the wild-type melatonin MT(1) receptor. Luzindole was subsequently confirmed as an antagonist of melatonin-mediated functional responses for both mutant receptors. These studies have identified that in the human melatonin MT(1) receptor, Ser(3.35) and Ser(3.39), in transmembrane domain 3, are critical for the formation of the high-affinity ligand binding site for agonists and partial agonists but not for the antagonist luzindole.
Conway
S
S
Molecular Neuroendocrinology Group, Division of Appetite and Energy Balance, Rowett Research Institute, Bucksburn, Aberdeen, AB21 9SB, United Kingdom. sco@rri.sari.ac.uk
Mowat
E S
ES
Drew
J E
JE
Barrett
P
P
Delagrange
P
P
Morgan
P J
PJ
D000081:Acetamides / Q000494:pharmacology; D000577:Amides / Q000494:pharmacology; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive / Q000187:drug effects; D019556:COS Cells; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008550:Melatonin / Q000031:analogs & derivatives / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D009281:Naphthalenes / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D044094:Receptors, Melatonin; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014162:Transfection; D014363:Tryptamines / Q000494:pharmacology
D000081:Acetamides; D000577:Amides; D007211:Indoles; D009281:Naphthalenes; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D044094:Receptors, Melatonin; D011993:Recombinant Fusion Proteins; C407519:S 20642; D014363:Tryptamines; C043464:N-acetyltryptamine; C057154:luzindole; C084711:agomelatine; D012694:Serine; C002465:methoxytryptophol; C044556:2-iodomelatonin; D000242:Cyclic AMP; D020962:Heterotrimeric GTP-Binding Proteins; D008550:Melatonin