A MUTATION CHANGES LIGAND SELECTIVITY AND TRANSMEMBRANE SIGNALING PREFERENCE OF THE NEUROKININ-1 RECEPTOR

studied the biochemical properties of a genetically engineered neuroki nin-1 receptor (NK(1)R) in which two residues lying on the extracellul ar edge of the fourth transmembrane domain were replaced by equivalent ly located elements of the neurokinin-2 receptor (G166C, Y167F NK(1)R mutant), The mutation produced two effects, The first is enhancement o f the apparent binding affinity for heterologous tachykinins (substanc e K and neurokinin B) and for N- or C-terminal modified analogues of s ubstance P, but not for substance P itself, its full-length analogues, and several peptide and nonpeptide antagonists, Only two antagonists, as exceptions, were found 60 exhibit a diminished affinity for the mu tant receptor. The second effect is a shift in NK(1)R preference for d istinct G protein-mediated signaling pathways, NK(1)R mediated phospho inositide hydrolysis was enhanced both in transiently and permanently transfected cells, while stimulation of cAMP accumulation did not chan ge in transient expression experiments and was reduced in permanently expressing cells. The effect of the mutation on ligand affinity was no t related to any obvious structural commonality, nor to the selectivit y for different neurokinin receptors or the agonistic/antagonistic nat ure of the ligand, However, all ligands responding to the mutation app ear to share the ability to induce phosphoinositide signaling more eff iciently than cAMP responses when binding to NK(1)R. We suggest that t he mutation shifts the internal equilibria of different functional for ms of NK(1)R. A theoretical analysis according to a multistate alloste ric model suggests that the link between binding and biological change s can result from altered stability constants of substates in the conf ormational space of the receptor.