Highly Selective Y₄ Receptor Antagonist Binds in an Allosteric Binding Pocket

Abstract

Human neuropeptide Y receptors (Y 1 R, Y 2 R, Y 4 R, and Y 5 R) belong to the superfamily of G protein-coupled receptors and play an important role in the regulation of food intake and energy metabolism. We identified and characterized the first selective Y 4 R allosteric antagonist (S)-VU0637120, an important step toward validating Y receptors as therapeutic targets for metabolic diseases. To obtain insight into the antagonistic mechanism of (S)-VU0637120, we conducted a variety of in vitro, ex vivo, and in silico studies. These studies revealed that (S)-VU0637120 selectively inhibits native Y 4 R function and binds in an allosteric site located below the binding pocket of the endogenous ligand pancreatic polypeptide in the core of the Y 4 R transmembrane domains. Taken together, our studies provide a first-of-its-kind tool for probing Y 4 R function and improve the general understanding of allosteric modulation, ultimately contributing to the rational development of allosteric modulators for peptide-activated G protein-coupled receptors (GPCRs).

Binding of Natural Peptide Ligands to the Neuropeptide Y5 Receptor

“…With respect to the N‐terminus of the ligand, our model does not predict direct interactions with the Y 5 R, similar to the NPY‐Y 2 R and PP‐Y 4 R models, [11–13] whereas the N‐terminus of NPY was suggested to interact extensively with Y 1 R [9] . Indeed, N‐terminally truncated NPY can bind Y 2 R but not Y 1 R [40] .…”

“…[11] Q 3.32 is highly conserved among the YRs and this interaction likely represents ah allmark for the entire multiligand/multireceptor system. [21] With respect to the N-terminus of the ligand, our model does not predict direct interactions with the Y 5 R, similar to the NPY-Y 2 Ra nd PP-Y 4 Rm odels, [11][12][13] whereas the Nterminus of NPY was suggested to interact extensively with Y 1 R. [9] Indeed, N-terminally truncated NPY can bind Y 2 Rbut not Y 1 R. [40] Interestingly,w hile Ala substitution the Nterminus of NPY barely affected Y 5 Ra ctivation, truncation of the 12 N-terminal residues reduced the potencyo ft he ligand by 8-fold. As altering the structure of the N-terminal region of NPY can heavily affect ligand potencye ven at YR subtypes that tolerate its truncation, [41] the N-terminal segment may stabilize the overall conformation of the peptide.…”

“…[9,10] Computational models of peptide-YR complexes have been proposed, based on experimental data obtained with different approaches:the NPY-Y 1 R, [9] theN PY-Y 2 R, [11] and the PP-Y 4 Rc omplex. [12,13] Some differences in the binding mode of Yp eptides have been identified.…”

“…Only the structures of the Y 1 R and most recently the Y 2 R, both in complex with small‐molecule antagonists, have been solved [9, 10] . Computational models of peptide‐YR complexes have been proposed, based on experimental data obtained with different approaches: the NPY‐Y 1 R, [9] the NPY‐Y 2 R, [11] and the PP‐Y 4 R complex [12, 13] . Some differences in the binding mode of Y peptides have been identified.…”

The Structural Basis of Peptide Binding at Class A G Protein-Coupled Receptors

“…Solution NMR studies showed a different structure with the helix present through the C-terminal end of the peptide [ 140 ]. The recent mutagenesis and docking study also suggested that the C-terminus of PP needed to unwind to bind to the Y 4 receptor [ 141 ]. Characterization of NPY in the membrane-bound state by NMR, CD, and EPR found the helix extending from residue 14 through the C-terminus [ 142 , 143 ].…”