Activation of the CB1 receptor is modulated by aspartate residue D2.63176 in transmembrane helix 2. Modeling studies identified residue K373 in the extracellular (EC)-3 loop in charged interactions with D2.63. To investigate this possibility, reciprocal mutations and biochemical studies were performed. None of the mutations resulted in a significant change in the binding affinity of SR141716A [N-(piperidiny-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride] or CP55,940 [(−)-cis-3-[2-hydroxyl-4-(1,1-dimethylheptyl)phenyl]-trans-4-[3-hydroxyl-propyl] cyclohexan-1-ol]. However, the functional signaling of CP55,940 and WIN55,212-2 [(R)-(+)-[2,3-dihydro-5-methyl-3-[(4-morpholinyl)methyl]-pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl](1-naphthalenyl)methanone mesylate] was impaired at the D2.63176A-K373A and single alanine mutants. The reciprocal D2.63176K-K373D mutant regained function for CP55,940 and WIN55,212-2. These results suggest that the ionic interactions between D2.63176 and K373 are important for CB1 signal transduction.
See article at J Pharmacol Exp Ther 2013, 345:189–197.
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