Direct and Differential Interaction of β-Arrestins with the Intracellular Domains of Different Opioid Receptors

Abstract
β-arrestins have been shown to play important roles in regulation of signaling and desensitization of opioid receptors in many in vivo studies. The current study was carried out to measure the direct interaction of β-arrestins with two functional intracellular domains, the third intracellular loop (I3L) and the carboxyl terminus (CT), of δ-, μ-, and κ-opioid receptors (DOR, MOR, and KOR, respectively). Results from the pull-down assay using glutathioneS-transferase fusion proteins demonstrated that β-arrestins (1 and 2) were able to bind to the I3L of DOR and to the CT of DOR and KOR. Surface plasmon resonance measurement gave similar results with typical dissociation equilibrium constant (KD) values in the micromolar range. The site-directed mutagenesis experiment further revealed that certain specific serine/threonine residues in these receptor domains play a critical role in their interaction with β-arrestins. Taken together, our data clearly indicated that β-arrestins interact differentially with the functional domains of different opioid receptors; this may provide a possible molecular basis for differential regulation of opioid receptors by β-arrestins.

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