The Leu/Val6.51 Side Chain of Cannabinoid Receptors Regulates the Binding Mode of the Alkyl Chain of Δ9-Tetrahydrocannabinol

Abstract

(-)-Δ⁹-trans-tetrahydrocannabinol (THC), which is the principal psychoactive constituent of Cannabis, mediates its action by binding to two members of the G-protein-coupled receptor (GPCR) family: the cannabinoid CB₁ (CB₁R) and CB₂ (CB₂R) receptors. Molecular dynamics simulations showed that the pentyl chain of THC could adopts an I-shape conformation, filling an intracellular cavity between Phe^3.36 and Trp^6.48 for initial agonist-induced receptor activation, in CB₁R but not in CB₂R. This cavity opens to the five-carbon chain of THC by the conformational change of the γ-branched, flexible, Leu^6.51 side chain of CB₁R, which is not feasible by the β-branched, mode rigid, Val^6.51 side chain of CB₂R. In agreement with our computational results, THC could not decrease the forskolin-induced cAMP levels in cells expressing mutant CB₁R^L6.51V receptor but could activate the mutant CB₂R^V6.51L receptor as efficiently as wild-type CB₁R. Additionally, JWH-133, a full CB₂R agonist, contains a branched dimethyl moiety in the ligand chain that bridges Phe^3.36 and Val^6.51 for receptor activation. In this case, the substitution of Val^6.51 to Leu in CB₂R makes JWH-133 unable to activate CB₂R^V6.51L. In conclusion, our combined computational and experimental results have shown that the amino acid at position 6.51 is a key additional player in the initial mechanism of activation of GPCRs that recognize signaling molecules derived from lipid species.