Hot topics: Agonist-bound crystal structures of the CB1 cannabinoid receptor

Antagonist bound crystal structures of GPCRs are useful in giving an insight into the molecular conformation of a receptor’s inactive state whilst enabling the design of new drugs. However, they prove insufficient to understand the activation mechanism of the receptor and mediation of its physiological effects. This necessitates the study of agonist-bound structures. In this direction, Hua et al., (2017) [1] have recently reported two agonist-bound crystal structures of Cannabinoid Receptor 1 (CB1), one with a tetrahydrocannabinol derivative, AM11542 [PDB: 5XRA], and the other with a hexahydrocannabinol, AM841 [PDB: 5XR8]. Previously, two antagonist-bound crystal structures of CB1 complexed with AM6538 and MK-0364 (taranabant) were reported by Hua et al., (2016) [PDB: 5TGZ] [2] and Shao et al., (2016) [PDB: 5U09], respectively [3].

Comparing the agonist and antagonist bound structures of the CB1 receptor reveals significant details:

1. The N-terminus in 5TGZ and 5U09 is a V-shaped loop which interacts with the bound antagonists, acting like a plug to the orthosteric binding pocket. The agonist-bound versions (5XRA and 5XR8), however, have their N-terminus residing over the binding pocket without any direct involvement in ligand binding. However, the N-terminus is truncated in all the crystal structures and hence the authors do not rule out the possibility that the full-length N-terminus might assume an entirely different conformation.

2. Both the agonists adopt an L-shaped conformation in the binding pocket, in contrast to the horizontal geometry of AM6538 in 5TGZ. The helical rearrangements hence observed in TM 1 and 2 and inward movement of residues Phe1702.57 and Phe1742.64 lead to a reduction in the binding pocket volume by 53% compared to 5TGZ. This serves as a testament to the highly flexible nature of the CB1 receptor and should be considered in future structure-based drug design studies for the receptor.

3. The alkyl chain of the two agonists extends into the ‘long channel’ of the receptor formed by the transmembrane helices (TM) 3, 5 and 6. The authors point out that this orientation is similar to that of ‘arm 2′- the nitroalkyl region of AM6538 in 5TGZ and of the alkyl chain of ML056 in the previously-described structure of the S1P1 receptor [4], thus indicating that the long channel could be a conserved binding region for alkyl chains in lipid binding receptors.

4. In 5TGZ and 5U09, the residues Phe2003.36 and Trp3566.48 exhibit aromatic stacking with each other. In this report, a synergistic conformational change of the residues was observed with the rotation of TM3 and side chain flip of Phe2003.36 towards the binding pocket occurring simultaneously with the rotation of TM6 away from TM3 breaking the interaction between the residues. The authors speculate the role of this ‘twin toggle switch’ in the activation of the receptor as a previous study has already shown [5].

5. Using the crystal structure, a cholesterol molecule has been identified to bind between the cytoplasmic portions of TM 2, 3, and 4 in the agonist-bound models. This was not observed in the antagonist models. However, the possible existence of a lipid access channel proposed in the taranabant bound (5U09) structure has not been discussed in this paper. This raises questions about the influence of lipids on the receptor binding through allosteric sites.

Comments by Lahari Murali (@wavesml), Steve Alexander (@mqzspa), Steven Doughty and Abi Emtage (@AbiEmtage)

[1] Hua, T. et al. (2017). Crystal structures of agonist-bound human cannabinoid receptor CB1. Nature.doi:10.1038/nature23272. [PMID: 28678776]

[2] Hua, T. et al. (2016). Crystal Structure of the Human Cannabinoid Receptor CB1. Cell 167: 750–762.e14. doi: 10.1016/j.cell.2016.10.004. [PMID: 27768894]

[3] Shao, Z. et al. (2016). High-resolution crystal structure of the human CB1 cannabinoid receptor. Nature 540:602–606. doi: 10.1038/nature20613. [PMID: 27851727]

[4] Hanson, M. A. et al. (2012). Crystal structure of a lipid G protein-coupled receptor. Science 335:851–855. doi: 10.1126/science.1215904. [PMID: 22344443]

[5] Singh, R., Hurst, D.P., Barnett-Norris, J., Lynch, D.L., Reggio, P.H., and Guarnieri, F. (2002). Activation of the cannabinoid CB1 receptor may involve a W6 48/F3 36 rotamer toggle switch. J. Pept. Res. 60: 357–370. [PMID: 12464114]

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