The gonadotrophin-releasing hormone (GnRH) receptor is unlike other G-protein coupled receptors in that the highly conserved amino acids, Asp in the second transmembrane region and Asn in the seventh, are interchanged. Site-directed mutagenesis studies mutated these residues back to their normally conserved positions. Two single mutants Asn87Asp & Asp318Asn and one double mutant Asn87Asp Asp318Asn were transiently expressed in COS-1 cells and their effect on binding to GnRH and inositol phosphate production measured. The single mutant Asp318Asn had no effect on ligand binding but abolished GnRH-dependent inositol phosphate production, whereas mutations Asn87Asp and Asn87Asp Asp318Asn show a complete loss of GnRH binding and subsequent inactivation of its second messenger system.