CONFORMATIONAL COMPATIBILITY AS A BASIS OF DIFFERENTIAL AFFINITIES OFTACHYKININS FOR THE NEUROKININ-1 RECEPTOR

The neurokinin-1 receptor is characterized by sub-nanomolar affinity f or substance P and 30-100 nM affinity for other substance P-related pe ptides, including neurokinin B and septide. We have characterized a ne urokinin-1 receptor mutant containing a Y216A substitution in the fift h transmembrane segment. This receptor mutant binds substance P with s ub-nanomolar affinity but loses much of its peptide discrimination cap ability, exhibiting 1-2 nM binding affinity for other tachykinins. Kin etic measurements of ligand binding indicate that the increased bindin g affinity of neurokinin B and septide for the Y216A mutant compared t o the wild-type receptor is due to a 100-fold increase in the associat ion rate constant without appreciable change in the dissociation rate constant. The substantially increased association rate constant for th e Y216A mutant suggests that the mutant receptor is probably more flex ible in accommodating the approaching peptide molecule. It is proposed that a major determinant of peptide specificity for the wild-type neu rokinin-1 receptor is the overall conformational compatibility between the receptor and the ligand, rather than residue-specific interaction s with the divergent N-terminal residues of different peptides. Furthe rmore, the highly conserved nature of Tyr-216 in the G protein coupled receptor family suggests that this residue may also play an important role in the receptor activation process in general.