Structure and function of serotonin G protein-coupled receptors

Abstract

Serotonin receptors are prevalent throughout the nervous system and the periphery, and remain one of the most lucrative and promising drug discovery targets for disorders ranging from migraine headaches to neuropsychiatric disorders such as schizophrenia and depression. There are 14 distinct serotonin receptors, of which 13 are G protein-coupled receptors (GPCRs), which are targets for approximately 40% of the approved medicines. Recent crystallographic and biochemical evidence has provided a converging understanding of the basic structure and functional mechanics of GPCR activation. Currently, two GPCR crystal structures exist for the serotonin family, the 5-HT1B and 5-HT2B receptor, with the antimigraine and valvulopathic drug ergotamine bound. The first serotonin crystal structures not only provide the first evidence of serotonin receptor topography but also provide mechanistic explanations into functional selectivity or biased agonism. This review will detail the findings of these crystal structures from a molecular and mutagenesis perspective for driving rational drug design for novel therapeutics incorporating biased signaling.

Keywords: 5-HT(1B); 5-HT(2B); Functional selectivity; GPCR; Serotonin; β-Arrestin.

Figure 1. 5-HT receptor ligands

Shown are the structures of 5-hydrotryptamine (5-HT or serotonin) and ergolines, lysergic acid diethylamide (LSD), ergotamine (ERG), and dihydroergotamine (DHE), which are ligands at all serotonin receptors. Also shown is an example triptan ligand, sumatriptan, which posseses 5-HT_1B receptor selectivity and also norfenfluramine, the active metabolite of fenfluramine, which possesses 5-HT_2B agonist activity.

Figure 2. ERG occupies both the orthosteric and extended binding sites in 5-HT_1B and 5-HT_2B receptors

Ergotamine (ERG) occupies two distinct sites in both the 5-HT_1B and 5-HT_2B receptors, the orthosteric site, where the indole nucleus of ERG resides, and the extended binding site, where the tripeptide portion is engaged.

Figure 3. Orthosteric sites of the 5-HT_1B and 5-HT_2B receptor crystal structures

Alignment of the 5-HT_1B (purple) and 5-HT_2B (blue) crystal structures reveals similar binding poses for ERG in the orthosteric binding pocket. A) Conserved aspartate (D3.32) forms a salt bride with protonated N⁶ of ERG and is also coordinated with residue 3.36. D3.32 also forms a hydrogen bond with Y7.43. B) Conserved residues, T3.37 and A5.46, engage the indole N-H portion of ERG. C) Aromatic residues F6.51, F6.52 and W6.48 involved in an edge-to-face π-π stacking with the ring system of ERG. D) Polar residues 5.43 and 6.55 involved in an interhelical hydrogen bond between TM5 and TM6, which are the putative residues for 5-hydroxy recognition of 5-HT. E) Summary of residue interactions with ERG in the orthosteric site.

Figure 4. Extended binding sites of the 5-HT_1B and 5-HT_2B receptor crystal structures

Alignment of the 5-HT_1B (purple) and 5-HT_2B (blue) crystal structures reveals similar yet strikingly different poses for ERG in the extended binding site of the binding pocket. A) Tripeptide portion of ERG forms similar interactions with nonpolar contacts in EL2 and residue 7.35 B) Rotamer state of tripeptide phenyl differs depending on presence of methionine at either 5.39 (5-HT_2B) or 6.58 (5-HT_1B) C) TM5 is pushed toward TM6 in 5-HT_2B receptor compared to the 5-HT_1B receptor, likely a result of the tripeptide phenyl rotamer state D) Summary of residue interactions with ERG in the extended binding site.

Figure 5. Microswitch trigger motifs of the 5-HT_1B and 5-HT_2B receptor crystal structures

The 5-HT_1B (purple) and 5-HT_2B (blue) crystal structures show select differences in their trigger motifs. A) Ionic lock between Asp3.49 and Arg3.50 in the DRY motif is broken in the 5-HT_1B structure but intact in the 5-HT_2B structure, suggestive of an inactive state. B) In the NPxxY motif, movement of Tyr7.53 toward the helical bundle is more pronounced in the 5-HT_2B structure compared to the 5-HT_1B structure. C) Inward movement of Phe6.44 in the P-I-F trigger motif is only present in the 5-HT_1B structure. D) Sodium (purple) in the sodium site in the DOR (orange) aligned with 5-HT structures show a condensed or collapsed sodium pocket in both 5-HT structures.

Figure 6. Tripeptide portion of ERG occupies a site overlapping with the allosteric modulator, LY2119620, in the muscarinic M2 receptor

Alignment of the 5-HT_2B (blue) and muscarinic M2 (green; PDB 4MQS) receptor crystal structures show that M2 orthosteric agonist iperoxo (IPX) and the ergoline nucleus of ERG occupy the same site deep in the helical bundle. Similary, the tripeptide portion of ERG and M2 allosteric modulator, LY2119620 (LY), also occupy the same site encompassing the extracellular portion of the receptor.