NMDA Receptor smFRET Studies Reveal Role of Dynamics of the Agonist-Binding Domain in Mediating Agonist Efficacy

Abstract

Ionotropic glutamate receptors are tetrameric ligand-gated ion channels which mediate the majority of excitatory neurotransmission in the central nervous system. This family is subdivided into three classes: the AMPA receptor, the kainate receptor, and the NMDA receptor. Extent of cleft closure of the agonist-binding domain is one mechanism by which the agonist mediates channel activity for a number of the glutamate receptor subtypes. However, only an open-cleft or a closed-cleft conformation has seen in crystal structures of the glycine-binding GluN1 subunit of the NMDA receptor, and no partially-closed cleft states have been observed. Here, we have used single molecule FRET to examine the dynamics of the NMDA receptor specifically with respect to the cleft closure conformational change of the isolated agonist-binding domain of GluN1 when bound to ligands of varying efficacy. These studies reveal differences in the range of cleft closure states occupied by the agonist-binding domain with the antagonist DCKA-bound form and the full agonist glycine-bound form showing a large range of cleft closure states, while the partial agonists ACBC and L-alanine, as well as the full agonist D-serine, have a much narrower spread in their cleft closure states. Further analysis shows that the fractional occupancy of the isolated domain in cleft-closure states below a threshold does correlate with agonist efficacy, providing a link between agonist-binding domain dynamics, along with cleft closure, and channel activity.