Crystal structure of the human NK1 tachykinin receptor

Abstract

The NK₁ tachykinin G-protein-coupled receptor (GPCR) binds substance P, the first neuropeptide to be discovered in mammals. Through activation of NK₁R, substance P modulates a wide variety of physiological and disease processes including nociception, inflammation, and depression. Human NK₁R (hNK₁R) modulators have shown promise in clinical trials for migraine, depression, and emesis. However, the only currently approved drugs targeting hNK₁R are inhibitors for chemotherapy-induced nausea and vomiting (CINV). To better understand the molecular basis of ligand recognition and selectivity, we solved the crystal structure of hNK₁R bound to the inhibitor L760735, a close analog of the drug aprepitant. Our crystal structure reveals the basis for antagonist interaction in the deep and narrow orthosteric pocket of the receptor. We used our structure as a template for computational docking and molecular-dynamics simulations to dissect the energetic importance of binding pocket interactions and model the binding of aprepitant. The structure of hNK₁R is a valuable tool in the further development of tachykinin receptor modulators for multiple clinical applications.

Keywords: GPCR; drug design; ligand recognition; substance P; tachykinin receptor.

Fig. 1.

Structural features of hNK₁R; (A) Global structure of the hNK₁R-PGS fusion protein from side view. The hNK₁R is represented as a green cartoon, with the PGS domain (wheat) fused between TMs 5 and 6. L760735 is shown as spheres with yellow carbons. (B) Extracellular view of hNK₁R, with PGS domain removed. (C) Surface representation as in B.

Fig. 2.

Stereoview of polder OMIT map (46) for the ligand L760735 (yellow sticks). The contour level is 3.1σ.

Fig. 3.

L760735 interaction with hNK₁R. (A) All residues within 4 Å of L760735 are depicted with cyan carbons. (B) Two-dimensional schematic of contacts between L760735 and hNK₁R, produced in Ligplot (65); (C) Chemical structures of aprepitant and L760735. (D, Top) Extracellular view, receptor surface is colored according to electrostatic potential. (D, Bottom) Side view, cut through the receptor.

Fig. 4.

Binding of substance P and L760735 to wild-type hNK₁R and point mutants. (A) Saturation binding isotherms for [³H]-substance P. (B) Competition binding assay for L760735, using [³H]-substance P as a probe. Error bars denote SE (n = 3 separate assays, each in duplicate).

Fig. 5.

Top flexible docking poses for L760735 and aprepitant in both neutral and protonated states. The L760735 ligand in the crystal structure is shown as orange sticks, while the docked ligands are shown as yellow sticks. Residues that form hydrogen bonds (dashed lines) with L760735 and aprepitant are shown as cyan sticks. (A) L760735 (neutral, ΔG_glide = −9.50, rmsd = 1.83 Å, best pose); (B) L760735 (charged, ΔG_glide = −9.37, rmsd = 1.73 Å, best pose); (C) aprepitant (neutral, ΔG_glide = −9.12, rmsd = 1.27 Å, best pose); (D) aprepitant (charged, ΔG_glide = −8.91, rmsd = 1.97 Å, second best pose). Glide docking scores are in kcal/mol.