ASN(102) OF THE GONADOTROPIN-RELEASING-HORMONE RECEPTOR IS A CRITICALDETERMINANT OF POTENCY FOR AGONISTS CONTAINING C-TERMINAL GLYCINAMIDE

We demonstrate a critical role for Asn(102) of the human gonadotropin- releasing hormone (GnRH) receptor in the binding of GnRH, Mutation of Asn(102), located at the top of the second transmembrane helix, to Ala resulted in a 225-fold loss of potency for GnRH. Eight GnRH analogs, all containing glycinamide C termini like GnRH, showed similar losses of potency between 95- and 750-foId for the [Ala(102)]GnRHR, compared with wildtype receptor, In contrast, four GnRH analogs that had ethyla mide in place of the C-terminal glycinamide residue, showed much small er decreases in potency between 2.4- and 11-fold, In comparisons of th ree agonist pairs, differing only at the C terminus, glycinamide deriv atives showed an 11-20-fold greater loss of potency for the mutant rec eptor than their respective ethylamide derivatives, Thus Asn(102) is a critical determinant of potency specifically for ligands with C termi nal glycinamide, while ligands with C-terminal ethylamide are less dep endent on Asn(102), These findings indicate a role for Asn(102) in the docking of the glycinamide C terminus and are consistent with hydroge n bonding of the Asn(102) side chain with the C-terminal amide moiety, Taken with previous data, they suggest a region of the GnRH receptor formed by the top of helices 2 and 7 as a binding pocket for the C-ter minal part of the ligand.