DELINEATION OF THE PEPTIDE BINDING-SITE OF THE HUMAN GALANIN RECEPTOR

Galanin, a neuroendocrine peptide of 29 amino acids, binds to G(i)/G(o )-coupled receptors to trigger cellular responses, To determine which amino acids of the recently cloned seven-transmembrane domain-type hum an galanin receptor are involved in the high-affinity binding of the e ndogenous peptide ligand, rye performed a mutagenesis study, Mutation of the His264 or His267 of transmembrane domain VI to alanine, or of P he282 of transmembrane domain VII to glycine, results in an apparent l oss of galanin binding, The substitution of Glu271 to serine in the ex tracellular loop III of the receptor causes a 12-fold loss in affinity for galanin, We combined the mutagenesis results with data on the pha rmacophores (Trp2, Tyr9) of galanin and with molecular modelling of th e receptor using bacteriorhodopsin as a model, Based on these studies, Re propose a binding site model for the endogenous peptide ligand in the galanin receptor where the N-terminus of galanin hydrogen bonds wi th Glu271 of the receptor, Trp2 of galanin interacts with the Zn2+ sen sitive pair of His264 and His267 of transmembrane domain Vf, and Tyr9 of galanin interacts with Phe282 of transmembrane domain VII, while th e C-terminus of galanin is pointing towards the N-terminus of the rece ptor.