Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2016 Oct 20;167(3):750-762.e14.
doi: 10.1016/j.cell.2016.10.004.

Crystal Structure of the Human Cannabinoid Receptor CB1

Tian Hua  1 , Kiran Vemuri  2 , Mengchen Pu  3 , Lu Qu  1 , Gye Won Han  4 , Yiran Wu  5 , Suwen Zhao  5 , Wenqing Shui  5 , Shanshan Li  5 , Anisha Korde  2 , Robert B Laprairie  6 , Edward L Stahl  6 , Jo-Hao Ho  6 , Nikolai Zvonok  2 , Han Zhou  2 , Irina Kufareva  7 , Beili Wu  8 , Qiang Zhao  8 , Michael A Hanson  9 , Laura M Bohn  10 , Alexandros Makriyannis  11 , Raymond C Stevens  12 , Zhi-Jie Liu  13
Affiliations

Crystal Structure of the Human Cannabinoid Receptor CB1

Tian Hua et al. Cell. .

Abstract

Cannabinoid receptor 1 (CB1) is the principal target of Δ9-tetrahydrocannabinol (THC), a psychoactive chemical from Cannabis sativa with a wide range of therapeutic applications and a long history of recreational use. CB1 is activated by endocannabinoids and is a promising therapeutic target for pain management, inflammation, obesity, and substance abuse disorders. Here, we present the 2.8 Å crystal structure of human CB1 in complex with AM6538, a stabilizing antagonist, synthesized and characterized for this structural study. The structure of the CB1-AM6538 complex reveals key features of the receptor and critical interactions for antagonist binding. In combination with functional studies and molecular modeling, the structure provides insight into the binding mode of naturally occurring CB1 ligands, such as THC, and synthetic cannabinoids. This enhances our understanding of the molecular basis for the physiological functions of CB1 and provides new opportunities for the design of next-generation CB1-targeting pharmaceuticals.

Keywords: AM6538; G protein-coupled receptor; THC; cannabinoid receptor CB1; cell signalling; crystal structure; marijuana; rimonabant; stabilizing antagonist.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Overall Structure of CB1-AM6538 Complex
(A) Side view of the CB1-AM6538 complex. The receptor is shown in gray cartoon representation. The ligand AM6538, shown with green sticks, demarcates the binding pocket which is partially occluded by the N-terminal loop (red). The nitrate group is not modeled in the experimental crystal structure as the electron density was not observed. The extracellular loops (ECLs) are shown in brown and the intracellular loops are shown in purple. (B) Top view of the extracellular side. The disulfide bond in ECL2 is shown as yellow sticks. See also Figures S2 and S3.
Figure 2
Figure 2. Synthesis and Characterization of AM6538
(A) Synthetic procedures for compound AM6538. (B) Saturation [3H]-CP55,940 binding assays in the absence (control) or presence of rimonabant (100 nM) or AM6538 (50 nM) demonstrates that both antagonists cause displacement of specific binding of the radioligand when present concurrently in the 1 h binding assay. (C) Following pretreatment of membranes (37 °C, 6 h) with buffer only (None), rimonabant (100 nM) or AM6538 (50 nM); membranes were washed with buffer 3× prior to [3H]-CP55,940 binding as described for B. (D) Percentage of remaining binding (Bmax) detected following the conditions described in (A) (concurrent) and (B) (pretreat & wash). Both antagonists decrease the binding of [3H]-CP55,940 to ~30% when incubated concurrently during the 1 h binding assay. Under pretreatment and washout conditions, rimonabant does not affect subsequent radioligand binding, while AM6538 continues to compete despite washing of the membranes. See also Figure S1 and Table S1.
Figure 3
Figure 3. Analysis of the Ligand Binding Pocket of CB1
(A) Key residues in CB1 for AM6538 binding. AM6538 (green carbons) and CB1 residues (teal carbons) involved in ligand binding are shown in stick representation. The receptor is shown in gray cartoon representation. (B) The shape of the ligand binding pocket. AM6538 (green carbons) and ML056 (brown carbons) are shown in stick representation. (C) Schematic representation of interactions between CB1 and AM6538. The 2,4-dichlorophenyl ring in the red circle is termed as arm 1; The 4-aliphatic chain substituted phenyl ring in the blue circle is termed as arm 2; The piperidin-1-ylcarbamoyl in the green circle is termed as arm 3. The nitrate group, which was not observed in the electron density, is shown in gray. (D) Electron density maps calculated from the refined structure of the CB1-AM6538 complex. |Fo|-|Fc| omit map (blue mesh) of the ligand AM6538 is shown (contoured at 3 σ). See also Figure S3.
Figure 4
Figure 4. Comparison of CB1, LPA1 and S1P1 Structural Features
(A) Side view of CB1 with structurally divergent regions of LPA1 (PDB ID: 4Z34) and S1P1 (PDB ID: 3V2Y) overlaid. LPA1 and S1P1 receptors are shown in gray cartoons. The CB1 N-terminal loop (red) occupies the polar binding pocket, helix I (red helix) is shifted out 7 Å compared with the other two receptors. ECL3 of CB1 shows a three helical turn extension of helix VII (brown helix). (B) 90° rotation of (A) for a top view of CB1 with structurally divergent regions of LPA1 (4Z34) and S1P1 (3V2Y) overlaid. CB1 shows a conserved conformation of ECL2 (blue) with the other two receptors, helix II is shifted out 2 Å (purple helix). (C–E) The interaction network of residues 3.28 of CB1 (K192), LPA1 (R124) and S1P1 (R120). Polar interactions are represented by black dashed lines. (C) CB1 is shown in gray cartoon, AM6538 is shown in green sticks and the key residues are shown in blue sticks; (D) LPA1 is shown in gray cartoon, ONO-9780307 is shown in purple-blue sticks and the key residues are shown in cyan sticks; (E) S1P1 is shown in gray cartoon, ML056 is shown in orange sticks and the key residues are shown in pink sticks. See also Figure S5.
Figure 5
Figure 5. Docking of Different Antagonists in the CB1 Crystal Structure
(A) CB1 binding pockets with rimonabant (blue sticks), otenabant (raspberry sticks) and taranabant (brown sticks) are shown in gray surface representation. (B) Chemical structures of rimonabant, otenabant and taranabant. The red/blue/green rectangles highlight previously described arms termed as arm 1/arm 2/arm 3 (see Figure 3C). (C) Predicted binding modes of rimonabant (blue sticks), otenabant (raspberry sticks) and taranabant (brown sticks) with CB1. The interacting residues are shown in yellow sticks and H178 is shown in green sticks. See also Figure S4 and Table S2.
Figure 6
Figure 6. Docking Poses of Different Cannabinoid Receptor Agonists
(A–F) Chemical structures and predicted binding poses of THC (pink sticks) (A), CP55,940 (magenta sticks) (B), Anandamide (cyan sticks) (C), 2-AG (green sticks) (D), JWH-018 (yellow sticks) (E), WIN 55,212-2 (blue sticks) (F). (G) Zoom-in view of predicted CP55,940 binding pose. CP55,940 is shown in magenta sticks, AM6538 is shown in green sticks and the key residues are shown in slate sticks. See also Figure S4 and Table S2.
See this image and copyright information in PMC

Comment in

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Abagyan R, Totrov M. Biased probability Monte Carlo conformational searches and electrostatic calculations for peptides and proteins. J Mol Biol. 1994;235:983–1002. - PubMed
    1. Abraham MJ, Murtola T, Schulz R, Páll S, Smith JC, Hess B, Lindahl E. GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX. 2015;1–2:19–25.
    1. Adams PD, Afonine PV, Bunkoczi G, Chen VB, Davis IW, Echols N, Headd JJ, Hung LW, Kapral GJ, Grosse-Kunstleve RW, et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Crystallogr. 2010;66:213–221. - PMC - PubMed
    1. Adams R, Mac KS, Jr, Loewe S. Tetrahydrocannabinol homologs with double branched alkyl groups in the 3-position. J Am Chem Soc. 1948;70:664–668. - PubMed
    1. Ahn KH, Bertalovitz AC, Mierke DF, Kendall DA. Dual role of the second extracellular loop of the cannabinoid receptor 1: ligand binding and receptor localization. Mol Pharmacol. 2009;76:833–842. - PMC - PubMed

Publication types

MeSH terms