Evaluating the DRY motif and its effects on ligand binding at the µ-Opioid Receptor

Opioids are considered the most efficacious drugs for management of moderate to severe pain, yet their use clinically is often restricted due to the onset of adverse side-effects. Drugs in this class produce most of their physiological effects (analgesia, nausea, vomiting) through activation of the μ-opioid receptor; however, an increasing number of studies have demonstrated that opioids can activate distinct downstream responses, a phenomenon termed functional selectivity. This project attempts to determine the amino acid residues involved in the activation of the μ-opioid receptor and the role they play in the binding of ligands to the receptor. The highly conserved DRY motif (aspartic acid-arginine-tyrosine) found commonly in GPCRs was mutated via site directed mutagenesis, aspartic acid-164 to alanine (A), glutamic acid (E), and arginine (R) and G-protein coupling and competition binding assays were performed to test whether ligand affinity and activity differed in the mutant receptors compared to the wild-type.