CB 1 cannabinoid receptor‐phosphorylated fourth intracellular loop structure‐function relationships

A peptide comprising the juxtamembrane C‐terminal intracellular loop 4 (IL4) of the CB 1 cannabinoid receptor possesses three serine residues (Ser402, Ser411, and Ser415). We report the effect of Ser phosphorylation on the CB 1 IL4 peptide conformation and cellular signaling functions using nuclear magnetic resonance spectroscopy, circular dichroism (CD), G protein activation, and cyclic adenosine monophosphate (cAMP) production. Phosphorylation at Ser residues induced helical structure in different environments. Helical content varies in the order of IL4p‐Ser411 > IL4pSer415 > IL4 > IL4pSer402. The efficacy of phosphorylated IL4 peptides in activating Go and Gi3 ([ 35 S]GTPγS binding) and inhibiting cAMP accumulation in N18TG2 cells was correlated with helicity changes. Bradykinin treatment, which activates protein kinase C (PKC), augmented CB 1 ‐mediated inhibition of cAMP accumulation, and this was reversed by a PKC inhibitor. We conclude that phosphorylation‐dependent alterations of helicity of CB 1 IL4 peptides can augment G protein signaling.