Sensing Allosteric Modulator Binding to AMPA Receptors at the Glutamate-Binding Site

Abstract

Positive allosteric modulators of AMPA receptors (GluAs) stabilize the activated GluA structure by binding to the glutamate-binding domain (LBD) dimer interface. By adding contacts across the dimer interface, allosteric modulators strengthen the LBD dimer and inhibit desensitization-linked dimer disruption. Structurally diverse molecules can occupy the interface to fill various pockets and act as LBD dimerizers. Functionally, allosteric modulators exert a primary allosteric effect by stabilizing the open channel; however, allosteric modulators with different dimer interface binding patterns lead to differing impacts on agonist site binding affinity. Here, we examine the secondary allosteric effects on the glutamate-binding site. Using nuclear magnetic resonance (NMR) spectroscopy, we observed the global backbone changes on the isolated GluA2 LBD as a result of allosteric modulator binding. Spectral changes at the dimer interface correlate well with allosteric modulator binding sites on bound X-ray crystal structures. The backbone residues near the agonist-binding site are also affected differently depending on the orientation of allosteric modulator binding. Further, allosteric modulator binding to the LBD induces local environmental changes at the agonist site as reported by NMR active nuclei on the LBD-bound agonist. By improving the molecular understanding of GluA allosteric modulators in the context of secondary agonist-site effects, our results provide further guidance for rational drug design.