An intracellular loop 2 amino acid residue determines differential binding of arrestin to the dopamine D2 and D3 receptors.

Lan H, Teeter MM, Gurevich VV, Neve KA
Mol Pharmacol. 2009 75 (1): 19-26

PMID: 18820126 · PMCID: PMC2606909 · DOI:10.1124/mol.108.050542

Dopamine D(2) and D(3) receptors are similar subtypes with distinct interactions with arrestins; the D(3) receptor mediates less agonist-induced translocation of arrestins than the D(2) receptor. The goals of this study were to compare nonphosphorylated arrestin-binding determinants in the second intracellular domain (IC2) of the D(2) and D(3) receptors to identify residues that contribute to the differential binding of arrestin to the subtypes. Arrestin 3 bound to glutathione transferase (GST) fusion proteins of the D(2) receptor IC2 more avidly than to the D(3) receptor IC2. Mutagenesis of the fusion proteins identified a residue at the C terminus of IC2, Lys149, that was important for the preferential binding of arrestin 3 to D(2)-IC2; arrestin binding to D(2)-IC2-K149C was greatly decreased compared with wild-type D(2)-IC2, whereas binding to the reciprocal mutant D(3)-IC2-C147K was enhanced compared with wild-type D(3)-IC2. Mutating this lysine in the full-length D(2) receptor to cysteine decreased the ability of the D(2) receptor to mediate agonist-induced arrestin 3 translocation to the membrane and decreased agonist-induced receptor internalization in human embryonic kidney 293 cells. The reciprocal mutation in the D(3) receptor increased receptor-mediated translocation of arrestin 3 without affecting agonist-induced receptor internalization. G protein-coupled receptor crystal structures suggest that Lys149, at the junction of IC2 and the fourth membrane-spanning helix, has intramolecular interactions that contribute to maintaining an inactive receptor state. It is suggested that the preferential agonist-induced binding of arrestin3 to the D(2) receptor over the D(3) receptor is due in part to Lys149, which could be exposed as a result of receptor activation.

MeSH Terms (22)

Amino Acid Sequence Amino Acid Substitution Animals Arrestin Binding Sites Biophysical Phenomena Cell Line Cysteine Glutathione Transferase Humans Hydrogen Bonding Kidney Models, Molecular Molecular Sequence Data Protein Binding Protein Structure, Secondary Protein Structure, Tertiary Rats Receptors, Dopamine D2 Receptors, Dopamine D3 Recombinant Fusion Proteins Sequence Homology, Amino Acid

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