TY - JOUR T1 - Modeling and Mutational Analysis of a Putative Sodium-Binding Pocket on the Dopamine D<sub>2</sub> Receptor JF - Molecular Pharmacology JO - Mol Pharmacol SP - 373 LP - 381 DO - 10.1124/mol.60.2.373 VL - 60 IS - 2 AU - Kim A. Neve AU - Medhane G. Cumbay AU - Kimberly R. Thompson AU - Rui Yang AU - David C. Buck AU - Val J. Watts AU - Curtiss J. DuRand AU - Martha M. Teeter Y1 - 2001/08/01 UR - http://molpharm.aspetjournals.org/content/60/2/373.abstract N2 - A homology model of the dopamine D2 receptor was constructed based on the crystal structure of rhodopsin. A putative sodium-binding pocket identified in an earlier model (PDB 1I15) was revised. It is now defined by Asn-419 backbone oxygen at the apex of a pyramid and Asp-80, Ser-121, Asn-419, and Ser-420 at each vertex of the planar base. Asn-423 stabilizes this pocket through hydrogen bonds to two of these residues. Highly conserved Asn-52 is positioned near the sodium pocket, where it hydrogen-bonds with Asp-80 and the backbone carbonyl of Ser-420. Mutation of three of these residues, Asn-52 in helix 1, Ser-121 in helix 3, and Ser-420 in helix 7, profoundly altered the properties of the receptor. Mutants in which Asn-52 was replaced with Ala or Leu or Ser-121 was replaced with Leu exhibited no detectable binding of radioligands, although receptor immunoreactivity in the membrane was similar to that in cells expressing the wild-type D2L receptor. A mutant in which Asn-52 was replaced with Gln, preserving hydrogen-bonding capability, was similar to D2L in affinity for ligands and ability to inhibit cAMP accumulation. Mutants in which either Ser-121 or Ser-420 was replaced with Ala or Asn had decreased affinity for agonists (Ser-121), but increased affinity for the antagonists haloperidol and clozapine. Interestingly, the affinity of these Ser-121 and Ser-420 mutants for substituted benzamide antagonists showed little or no dependence on sodium, consistent with our hypothesis that Ser-121 and Ser-420 contribute to the formation of a sodium-binding pocket. ER -