RT Journal Article SR Electronic T1 Molecular determinants for competitive inhibition of α4β2 nicotinic acetylcholine receptors JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP mol.110.065490 DO 10.1124/mol.110.065490 A1 Patricio Ernesto Iturriaga-Vasquez A1 Analisa Carbone A1 Olimpo Garcia-Beltran A1 Phil D Livingstone A1 Philip C Biggin A1 Bruce K Cassels A1 Susan Wonnacott A1 Gerald Zapata-Torres A1 Bermudez Isabel YR 2010 UL http://molpharm.aspetjournals.org/content/early/2010/06/14/mol.110.065490.abstract AB The Erythrina alkaloids erysodine and dihydro-β-erythroidine (DHβE) are potent and selective competitive inhibitors of α4β2 nicotinic acetylcholine receptors (nAChRs) but little is known about the molecular determinants of sensitivity of this receptor subtype to inhibition by this class of antagonists. We addressed this issue by examining the effects of DHβE and a range of aromatic Erythrina alkaloids on [3H]cytisine binding and receptor function in conjunction with homology models of the α4β2 nAChR, mutagenesis and functional assays. The lactone group of DHβE and a hydroxyl group at position C16 in aromatic Erythrina alkaloids were identified as major determinants of potency, which was decreased when the conserved residue Y126 in loop A of the α4 subunit was substituted by alanine. Sensitivity to inhibition was also decreased by substituting the conserved aromatic residues α4W182 (loop B), α4Y230 (loop C) and β2W82 (loop D) and the non-conserved β2T84, however only α4W182 was predicted to contact bound antagonist, suggesting α4Y230, β2W82 and β2T84 contribute allosterically to the closed state elicited by bound antagonist. Additionally, homology modelling predicted strong ionic interactions between the ammonium centre of the Erythrina alkaloids and β2D196, leading to uncapping of loop C. Consistent with this, β2D196A abolished sensitivity to inhibition by DHβE or erysodine but not by epierythratidine, which is not predicted to form ionic bonds with β2D196. This residue is not conserved in subunits that comprise nAChRs with low sensitivity to inhibition by DHβE or erysodine, which highlights β2D196 as a major determinant of the receptor selectivity of Erythrina alkaloids.The American Society for Pharmacology and Experimental Therapeutics