Asynchronous Movements Prior to Pore Opening in NMDA Receptors

Abstract

Glutamate-gated ion channels embedded within the neuronal membrane are the primary mediators of fast excitatory synaptic transmission in the CNS. The ion channel of these glutamate receptors contains a pore-lining transmembrane M3 helix surrounded by peripheral M1 and M4 helices. In the NMDA receptor subtype, opening of the ion channel pore, mediated by displacement of the M3 helices away from the central pore axis, occurs in a highly concerted fashion, but the associated temporal movements of the peripheral helices are unknown. To address the gating dynamics of the peripheral helices, we constrained the relative movements of the linkers that connect these helices to the ligand-binding domain using engineered cross-links, either within (intra-GluN1 or GluN2A) or between subunits. Constraining the peripheral linkers in any manner dramatically curtailed channel opening, highlighting the requirement for rearrangements of these peripheral structural elements for efficient gating to occur. However, the magnitude of this gating effect depended on the specific subunit being constrained, with the most dramatic effects occurring when the constraint was between subunits. Based on kinetic and thermodynamic analysis, our results suggest an asynchrony in the displacement of the peripheral linkers during the conformational and energetic changes leading to pore opening. Initially there are large-scale rearrangements occurring between the four subunits. Subsequently, rearrangements occur within individual subunits, mainly GluN2A, leading up to or in concert with pore opening. Thus, the conformational changes induced by agonist binding in NMDA receptors converge asynchronously to permit pore opening.

Block of NMDA receptor channels by endogenous neurosteroids: implications for the agonist induced conformational states of the channel vestibule

“…The compact geometry allows an optimal distance to form vdW contacts as well as hydrogen bonds among the side-chain atoms of threonine residues, and as the distance between the helices grows further, nonsymmetric opening/gating at these threonine residues progresses. These data agree well with the results of experiments in which the cysteine accessibility method and helix cross-linking were used to show the asymmetric contribution of the M3 segments from the GluN1 and GluN2 subunits to the opening of the extracellular NMDAR channel vestibule2638. In contrast to the WT results, in mutants, the non-symmetric behavior and the reduction of pore diameter in the gating region during the initial steps of opening are less prominent (Fig.…”

Structural modeling for the open state of an NMDA receptor

“…Several GluN1 residues, including F552, V642, L649, and L655, are outliers in the correlation plot. Future experimental studies will allow for more stringent tests of the open model, e.g., by designing cysteine substitutions that are predicted to form metal coordination or disulfide cross-linking (Heymann et al, 2013; Kazi et al, 2013). …”

Cysteine Substitution of Transmembrane Domain Amino Acids Alters the Ethanol Inhibition of GluN1/GluN2A N-Methyl-d-Aspartate Receptors

“…Whereas previous studies focused on a role for TM3 and TM4 residues in ethanol action (Ronald et al, 2001;Ren et al, 2003Ren et al, , 2012Honse et al, 2004;Smothers and Woodward, 2006;Xu et al, 2012), the present study is the first to show that TM1 residues may also contribute. Residues in extracellular regions link GluN1 TM1 to ligand-binding domains that interact with similar linker sites in GluN2A TM4 and may be involved in early steps of receptor activation that lead to TM3 domain movement (Talukder et al, 2010;Kazi et al, 2013). As TM3 is critically involved in channel gating (Jones et al, 2002;Chang and Kuo, 2008), ethanol perturbation of TM1 residues that interact with TM3 may alter the transitions between closed and open states of the receptor.…”