The role of the N-terminal and mid-region residues of substance P in regulating functional selectivity at the tachykinin NK1 receptor

Previous studies have shown that tachykinin peptide ligands of the tachykinin NK1 receptor exhibit functional selectivity with respect to signal activation and desensitization. The differences are most dramatic between the naturally occurring peptides substance P (RPKPQQFFGLM-NH2) and ranatachykinin C (HNPASFIGLM-NH2). To understand the structural features of the peptides that underlie these differences, four peptide analogs have been designed and tested. The analogs were designed to assess the major structural differences between substance P and ranatachykinin C, including the role of the N-terminal Arg and the substitution of the mid-region Glns with Ala and Ser (Q5 replaced with A and/or Q6 replaced with S). Receptor binding, receptor activation of intracellular calcium fluxes, and receptor desensitization of the rat tachykinin NK1 receptor were quantified for each ligand. All of the peptides bound to the rat tachykinin NK1 receptor with high affinity, produced robust calcium signal activation, and led to agonist-induced receptor desensitization. It was found that deletion of the N-terminal Arg of substance P or replacement of either or both Q5 and Q6 altered the functional selectivity of substance P based on the relationship of receptor binding to receptor activation and activation to desensitization. When considered in light of our previously published nuclear magnetic resonance structure data, the data presented herein suggest that the one, five and six positions of the substance P backbone are key structural residues that govern the relative degree of tachykinin peptide-mediated receptor signaling and desensitization.