Conformational analysis of the C-terminal Gly-Leu-Met-NH2 tripeptide of substance P bound to the NK-1 receptor

We examined the effect of simultaneously incorporating proline or proline-amino acid chimeras in positions 9, 10, and/or 11 of substance P, on the affinity for the two NK-1 binding sites and on second-messenger activation. Because these 3-substituted prolines constrain not only the (phi,psi) values of the peptide backbone, but also the X space of the amino acid side chain, we were able to gather data on the structural requirements for high-affinity binding to the NK-1 receptor. We were able to confirm that this C-terminal component is crucial and that it should adopt an extended conformation close to a polyproline II structure when bound to the receptor. The partial additivity of these constraints, more specifically, for the NK-1 M site, suggests that the peptide backbone flexibility around the hinge-point residue Gly9 is essential to subtly position crucial side chains.