Dataset Information

Subtype-dependent N-methyl-D-aspartate receptor amino-terminal domain conformations and modulation by spermine.

ABSTRACT: The N-methyl-d-aspartate (NMDA) subtype of the ionotropic glutamate receptors is the primary mediator of calcium-permeable excitatory neurotransmission in the central nervous system. Subunit composition and binding of allosteric modulators to the amino-terminal domain determine the open probability of the channel. By using luminescence resonance energy transfer with functional receptors expressed in CHO cells, we show that the cleft of the amino-terminal domain of the GluN2B subunit, which has a lower channel open probability, is on average more closed than the GluN2A subunit, which has a higher open probability. Furthermore, the GluN1 amino-terminal domain adopts a more open conformation when coassembled with GluN2A than with GluN2B. Binding of spermine, an allosteric potentiator, opens the amino-terminal domain cleft of both the GluN2B subunit and the adjacent GluN1 subunit. These studies provide direct structural evidence that the inherent conformations of the amino-terminal domains vary based on the subunit and match the reported open probabilities for the receptor.

SUBMITTER: Sirrieh RE

PROVIDER: S-EPMC4432297 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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Publications

Subtype-dependent N-methyl-D-aspartate receptor amino-terminal domain conformations and modulation by spermine.

Sirrieh Rita E RE MacLean David M DM Jayaraman Vasanthi V

The Journal of biological chemistry 20150331 20

The N-methyl-d-aspartate (NMDA) subtype of the ionotropic glutamate receptors is the primary mediator of calcium-permeable excitatory neurotransmission in the central nervous system. Subunit composition and binding of allosteric modulators to the amino-terminal domain determine the open probability of the channel. By using luminescence resonance energy transfer with functional receptors expressed in CHO cells, we show that the cleft of the amino-terminal domain of the GluN2B subunit, which has a ...[more]

PMID: 25829490

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Project description:Electrical synapses between neurons in the mammalian CNS are predominantly formed of the connexin36 (Cx36) gap junction (GJ) channel protein. Unique among GJs formed of a number of other members of the Cx gene family, Cx36 GJs possess a high sensitivity to intracellular Mg2+ that can robustly act to modulate the strength of electrical synaptic transmission. Although a putative Mg2+ binding site was previously identified to reside in the aqueous pore in the first extracellular (E1) loop domain, the involvement of the N-terminal (NT) domain in the atypical response of Cx36 GJs to pH was shown to depend on intracellular levels of Mg2+. In this study, we examined the impact of amino acid substitutions in the NT domain on Mg2+ modulation of Cx36 GJs, focusing on positions predicted to line the pore funnel, which constitutes the cytoplasmic entrance of the channel pore. We find that charge substitutions at the 8th, 13th, and 18th positions had pronounced effects on Mg2+ sensitivity, particularly at position 13 at which an A13K substitution completely abolished sensitivity to Mg2+. To assess potential mechanisms of Mg2+ action, we constructed and tested a series of mathematical models that took into account gating of the component hemichannels in a Cx36 GJ channel as well as Mg2+ binding to each hemichannel in open and/or closed states. Simultaneous model fitting of measurements of junctional conductance, gj, and transjunctional Mg2+ fluxes using a fluorescent Mg2+ indicator suggested that the most viable mechanism for Cx36 regulation by Mg2+ entails the binding of Mg2+ to and subsequent stabilization of the closed state in each hemichannel. Reduced permeability to Mg2+ was also evident, particularly for the A13K substitution, but homology modeling of all charge-substituted NT variants showed only a moderate correlation between a reduction in the negative electrostatic potential and a reduction in the permeability to Mg2+ ions. Given the reported role of the E1 domain in Mg2+ binding together with the impact of NT substitutions on gating and the apparent state-dependence of Mg2+ binding, this study suggests that the NT domain can be an integral part of Mg2+ modulation of Cx36 GJs likely through the coupling of conformational changes between NT and E1 domains.

Dataset Information

Subtype-dependent N-methyl-D-aspartate receptor amino-terminal domain conformations and modulation by spermine.

Publications

Subtype-dependent N-methyl-D-aspartate receptor amino-terminal domain conformations and modulation by spermine.

Similar Datasets

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