GLY(166) IN THE NK1 RECEPTOR REGULATES TACHYKININ SELECTIVITY AND RECEPTOR CONFORMATION

We have studied the pharmacological properties of genetically engineer ed human NK1 tachykinin receptors in which residues at the extracellul ar surface of the fourth transmembranal domain were substituted with t he corresponding amino acids from the NK2 receptor, We show that subst itution of G166C:Y167F in the human NK1 receptor induces high affinity binding of a group of tachykinin ligands, known as 'septides' (i.e. n eurokinin A, neurokinin B, [pGlu(6),Pro(9)]-substance P6-11 and substa nce P-methylester), In contrast, binding of substance P and non-peptid e antagonists is unaffected by these mutations, This effect parallels that found on the rat receptor and is therefore species specific, Seco nd, we demonstrate that mutation of Gly(166) to Cys alone is both nece ssary and sufficient to create this pan-reactive tachykinin receptor, whereas replacement of Tyr(167) by Phe has no detectable effect on the pharmacological properties of the receptor, Furthermore, analysis of the effect of N-ethylmaleimide and dithiothreitol on binding of radiol abelled substance P documents differences in the mode in which this li gand interacts with wild-type and mutant receptors and supports the ex istence of a mutational induced change in the conformational status of the NK1 receptor. (C) 1997 Federation of European Biochemical Societi es.