Abstract
Sodium ions (Na+) allosterically modulate the binding of orthosteric agonists and antagonists to many class A G protein-coupled receptors, including the dopamine D2 receptor (D2R). Experimental and computational evidences have revealed that this effect is mediated by the binding of Na+ to a conserved site located beneath the orthosteric binding site (OBS). SB269652 acts as a negative allosteric modulator (NAM) of the D2R that adopts an extended bitopic pose, in which the tetrahydroisoquinoline moiety interacts with the OBS and the indole-2-carboxamide moiety occupies a secondary binding pocket (SBP). In this study, we find that the presence of a Na+ within the conserved Na+-binding pocket is required for the action of SB269652. Using fragments of SB269652 and novel full-length analogues, we show that Na+ is required for the high affinity binding of the tetrahydroisoquinoline moiety within the OBS, and that the interaction of the indole-2-carboxamide moiety with the SBP determines the degree of Na+-sensitivity. Thus, we extend our understanding of the mode of action of this novel class of NAM by showing it acts synergistically with Na+ to modulate the binding of orthosteric ligands at the D2R, providing opportunities for fine-tuning of modulatory effects in future allosteric drug design efforts.
Highlights
Sodium ions (Na+) allosterically modulate the binding of orthosteric agonists and antagonists to many class A G protein-coupled receptors, including the dopamine D2 receptor (D2R)
Our previous study revealed that Glu952.65 within the secondary binding pocket (SBP) of the D2R was an important determinant of affinity and cooperativity of SB269652
The potentially conserved functional role of Na+ at GPCRs30 prompted us to explore the impact of Na+ binding on the pharmacology of SB269652, the first small-molecule negative allosteric modulator (NAM) identified at the D2R
Summary
Sodium ions (Na+) allosterically modulate the binding of orthosteric agonists and antagonists to many class A G protein-coupled receptors, including the dopamine D2 receptor (D2R). SB269652 acts as a negative allosteric modulator (NAM) of the D2R that adopts an extended bitopic pose, in which the tetrahydroisoquinoline moiety interacts with the OBS and the indole-2-carboxamide moiety occupies a secondary binding pocket (SBP). Using MD simulations, Selent and coworkers proposed that Na+ enters the D2R via the extracellular side of the receptor, moving along a network of negatively charged residues including Glu952.65 before entering the Na+-binding site coordinated by Asp802.50 This Na+ binding was proposed to lock the rotamer toggle switch Trp3866.48 in the inactive state[26]
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