Structural Proof of a Dimeric Positive Modulator Bridging Two Identical AMPA Receptor-Binding Sites

Abstract

Dimeric positive allosteric modulators of ionotropic glutamate receptors were designed, synthesized, and characterized pharmacologically in electrophysiological experiments. The designed compounds are dimers of arylpropylsulfonamides and have been constructed without a linker. The monomeric arylpropylsulfonamides were derived from known modulators and target the cyclothiazide-binding site at the AMPA receptors. The three stereoisomers--R,R, meso, and S,S--of the two constructed dimers were prepared, and in vitro testing showed the R,R forms to be the most potent stereoisomers. The biarylpropylsulfonamides have dramatically increased potencies, more than three orders of magnitude higher than the corresponding monomers. Dimer (R,R)-2a was cocrystallized with the GluR2-S1S2J construct, and an X-ray crystallographic analysis showed (R,R)-2a to bridge two identical binding pockets on two neighboring GluR2 subunits. Thus, this is biostructural evidence of a homomeric dimer bridging two identical receptor-binding sites.

Kinetic Contributions to Gating by Interactions Unique to N-methyl-d-aspartate (NMDA) Receptors

“…Based on these observations it was proposed that these mutations modulate current deactivation by changing the glutamate dissociation rate, although the exact mechanism is unknown. Consistent with this hypothesis, fast deactivating AMPA and kainate receptors lack this critical tyrosine residue but instead accommodate at this site allosteric modulators that also decrease deactivation (11,30,31). Therefore, it was proposed that the tyrosine residue uniquely present in NMDA receptors acts as a "natural" break in deactivation and renders NMDA receptors insensitive to AMPA receptor-specific positive modulators.…”

“…Site II Side-chain Truncations Impart Sensitivity to AMPA Receptor Positive Modulators-In AMPA receptors, the equivalent location of site II residues defines the binding pocket for small allosteric modulators, which potentiate currents by slowing macroscopic deactivation (11,30,31,(43)(44)(45)(46). The "floor" of this binding pocket is lined by serine residues absent in NMDA receptors (Fig.…”

Structure and Dynamics of AMPA Receptor GluA2 in Resting, Pre-Open, and Desensitized States

“…Because AMPA receptors desensitize rapidly and profoundly, crystallization in the presence of full or partial agonists should yield structures representing a desensitized receptor conformation. To trap the receptor in an agonist-bound, activated state, we employed agonists in combination with a high affinity positive allosteric modulator (R,R)-2b which block desensitizations (Kaae et al, 2007). Finally, crystallization under ligand-free conditions was carried out to determine a structure of the full-length GluA2 receptor in an apo/resting state.…”

X-ray structures of AMPA receptor–cone snail toxin complexes illuminate activation mechanism

“…Here we report x-ray crystal structures of an intact, rat GluA2 AMPA receptor in complex with a homodimeric Conus striatus cone snail toxin, con-ikot-ikot (8), with (R,R)-2b, a high affinity, 2-fold symmetric positive allosteric modulator (9), and with the partial agonist kainate (10) or the potent partial agonist fluorowillardiine (11). To do this, we expressed, purified and solved the structure of con-ikot-ikot, a disulfide-bond-rich polypeptide, previously shown to induce paralysis in fish and potently and selectively block desensitization of AMPA receptors (8) (Fig.…”