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Review
. 2020 Apr;287(8):1496-1510.
doi: 10.1111/febs.15128. Epub 2019 Nov 23.

Structural insights into melatonin receptors

Benjamin Stauch  1   2 , Linda C Johansson  1   2 , Vadim Cherezov  1   2   3
Affiliations
Review

Structural insights into melatonin receptors

Benjamin Stauch et al. FEBS J. 2020 Apr.

Abstract

The long-anticipated high-resolution structures of the human melatonin G protein-coupled receptors MT1 and MT2 , involved in establishing and maintaining circadian rhythm, were obtained in complex with two melatonin analogs and two approved anti-insomnia and antidepression drugs using X-ray free-electron laser serial femtosecond crystallography. The structures shed light on the overall conformation and unusual structural features of melatonin receptors, as well as their ligand binding sites and the melatonergic pharmacophore, thereby providing insights into receptor subtype selectivity. The structures revealed an occluded orthosteric ligand binding site with a membrane-buried channel for ligand entry in both receptors, and an additional putative ligand entry path in MT2 from the extracellular side. This unexpected ligand entry mode contributes to facilitating the high specificity with which melatonin receptors bind their cognate ligand and exclude structurally similar molecules such as serotonin, the biosynthetic precursor of melatonin. Finally, the MT2 structure allowed accurate mapping of type 2 diabetes-related single-nucleotide polymorphisms, where a clustering of residues in helices I and II on the protein-membrane interface was observed which could potentially influence receptor oligomerization. The role of receptor oligomerization is further discussed in light of the differential interaction of MT1 and MT2 with GPR50, a regulatory melatonin coreceptor. The melatonin receptor structures will facilitate design of selective tool compounds to further dissect the specific physiological function of each receptor subtype as well as provide a structural basis for next-generation sleeping aids and other drugs targeting these receptors with higher specificity and fewer side effects.

Keywords: G protein-coupled receptors; X-ray free-electron laser; chronobiology; diabetes; melatonin; serial femtosecond crystallography; serotonin; sleeping aids.

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Conflict of interest statement

Conflict of interest

The authors declare no competing interests.

Figures

Figure 1 |
Figure 1 |. Melatonin biosynthesis.
Serotonin is converted to melatonin in the pineal gland of the animal brain via two subsequent enzymatic reactions. The first step, N-acetylation, is catalyzed by serotonin N-acetyltransferase (SNAT), and inhibited by blue wavelength light signals transmitted from the retina via the suprachiasmic nucleus (SCN) resulting in a high level production of melatonin during the night and low level during the day. Melatonin is then released to the third ventricle and to the blood stream, allowing it to act on both brain and peripheral tissues where MT1 and MT2 are expressed. Among the diverse physiological functions of melatonin are regulation of body temperature and blood pressure, facilitating sleep.
Figure 2 |
Figure 2 |. Crystallographic packing of MT1.
The receptor was crystallized in tetragonal space group P4 21 2 with unit cell (red) dimensions of a=b=122.3 Å and c=122.8 Å (PDB identifier 6me2). Symmetry mates were found to pack in alternating layers mediated by head-to-head receptor (light blue, teal) weak contacts and strong contacts mediated by intracellular fusion proteins (dark blue, green). This figure has been generated using PyMOL.
Figure 3 |
Figure 3 |. Differences and similarities of MT receptor binding sites.
A, Ligand-binding sites of 2-pmt-bound MT1 (light blue, left; PDB identifier 6me3) and MT2 (light orange, right; PDB 6me6) receptors are shown as closed surfaces with receptors in cartoon representation. 2-pmt (green) as well as the three key ligand-coordinating residues and the disulfide bridge are shown as sticks with blue nitrogens, red oxygens, and yellow sulfurs. Hydrogen bonds are indicated by yellow dashed lines. Amino-acid sequence and conformations within the binding site are remarkably conserved between receptor subtypes; the most apparent differences include the smaller volume in MT1 (710 Å3) compared to MT2 (766 Å3), mostly due to a small compaction around the R2 tail of the ligand (orange dash curve with a black arrow), as well as narrowing of the lateral channel (dashed black arrow) and expansion of the ECL access in MT2 (solid black arrows). B, position of putative alcohol (magenta) close to residue N255 in binding site of MT1. See Figure 3C for definition of melatonin substituents R1, R2, and R3. C, chemical structure of melatonin. Heteroatom numbering based on the indole numbering-scheme are indicated as blue digits, and positions with variable substituents in melatonin derivatives are indicated by R1, R2, and R3. This figure has been generated using PyMOL.
Figure 4 |
Figure 4 |. Extracellular occlusion in class A GPCRs.
A, surface section of melatonin receptors MT1 (left) and MT2 (middle) compared to serotonin receptor 5-HT2C (right, PDB identifier 6bqg). Lipid bilayer indicated as light grey schematic. Ligand access routes to respective receptors are indicated by blue arrows. B, conformation of extracellular parts of MT1 (left and first insert; PDB 6me2) compared to rhodopsin (PDB 1u19) and lipid receptors (prostaglandin receptors EP3 and EP4, PDB identifiers 6m9t and 5ywy; sphingosine-1-phospate receptor S1P1, PDB 3v2w; lysophosphatidic acid receptor LPA1, PDB 4z35; cannabinoid receptors CB1 and CB2, PDBs 5xr8 and 5zty) shown as inserts (N-terminus, green; ECL2, magenta; disulfides, sticks with yellow sulfurs). Ligands are shown as spheres with blue nitrogens and red oxygens, unresolved non-terminal extracellular residues as dashed lines, and helices as cylinders. Rotation angles are approximate. This figure has been generated using PyMOL.
Figure 5 |
Figure 5 |. Location of T2D-related single nucleotide polymorphisms (SNPs) on the MT2 structure.
Receptor shown in cartoon representation with solvent-accessible surface (PDB identifier 6me6). Ligand 2-pmt shown as spheres, and SNPs shown as sticks with blue nitrogens and red oxygens. SNPs located in the TM1-TM2 region of the receptor (P36, A42, A52, L60, and P95) and potentially involved in receptor oligomerization or interactions with other membrane partners are colored cyan, while SNPs clustered in the intracellular region responsible for binding transducers are colored magenta, or green otherwise. This figure has been generated using PyMOL.
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