An inter-dimer allosteric switch controls NMDA receptor activity

NMDA receptors (NMDARs) are glutamate-gated ion channels that are key mediators of excitatory neurotransmission and synaptic plasticity throughout the central nervous system. They form massive heterotetrameric complexes endowed with unique allosteric capacity provided by eight extracellular clamshell-like domains arranged as two superimposed layers. Despite an increasing number of full-length NMDAR structures, how these domains cooperate in an intact receptor to control its activity remains poorly understood. Here, combining single-molecule and macroscopic electrophysiological recordings, cysteine biochemistry, and in silico analysis, we identify a rolling motion at a yet unexplored interface between the two constitute dimers in the agonist-binding domain (ABD) layer as a key structural determinant in NMDAR activation and allosteric modulation. This rotation acts as a gating switch that tunes channel opening depending on the conformation of the membrane-distal N-terminal domain (NTD) layer. Remarkably, receptors locked in a rolled state display “super-activity” and resistance to NTD-mediated allosteric modulators. Our work unveils how NMDAR domains move in a concerted manner to transduce long-range conformational changes between layers and command receptor channel activity.