Species selectivity of nonpeptide antagonists of the gonadotropin-releasing hormone receptor is determined by residues in extracellular loops II and III and the amino terminus.
Efforts to develop orally available gonadotropin-releasing hormone (GnRH) receptor antagonists have led to the discovery of several classes of potent nonpeptide antagonists. Here we investigated molecular interactions of three classes of nonpeptide antagonists with human, rat, and macaque GnRH receptors. Although all are high affinity ligands of the human receptor (K(i) <5 nm), these compounds show reduced affinity for the macaque receptor and bind only weakly (K(i) >1 microm) to the rat receptor. To identify residues responsible for this selectivity, a series of chimeric receptors and mutant receptors was constructed and evaluated for nonpeptide binding. Surprisingly, 4 key residues located in the amino terminus (Met-24) and extracellular loops II (Ser-203, Gln-208) and III (Leu-300) of the GnRH receptor appear to be primarily responsible for species-selective binding. Comparisons of reciprocal mutations suggest that these may not be direct contacts but rather may be involved in organizing extracellular portions of the receptor. These data are novel because most previous reports of residues involved in binding of nonpeptide ligands to peptide-activated G protein-coupled receptors, including the GnRH receptor as well as mono-amine receptors, have identified binding sites in the transmembrane regions.
Reinhart
Greg J
GJ
Department of Endocrinology, Neurocrine Biosciences Inc., 10555 Science Center Drive, San Diego, CA 92121, USA.
Xie
Qiu
Q
Liu
Xin-Jun
XJ
Zhu
Yun-Fei
YF
Fan
Jun
J
Chen
Chen
C
Struthers
R Scott
RS
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D016466:CHO Cells; D002417:Cattle; D002678:Chimera; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008024:Ligands; D008253:Macaca mulatta; D008956:Models, Chemical; D009154:Mutation; D010446:Peptide Fragments / Q000494:pharmacology; D010455:Peptides / Q000737:chemistry; D017354:Point Mutation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D011966:Receptors, LHRH / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D012243:Rhodopsin / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014162:Transfection
D008024:Ligands; D010446:Peptide Fragments; D010455:Peptides; D011966:Receptors, LHRH; D012243:Rhodopsin