Residues at the tip of the pore loop of NR3B-containing NMDA receptors determine Ca2+ permeability and Mg2+ block
1 Department of Biochemistry I - Receptor Biochemistry, Ruhr University Bochum, Universitaetsstrasse 150, D-44780 Bochum, Germany
2 International Graduate School of Neuroscience (IGSN), Ruhr University Bochum, Bochum, Germany
3 Ruhr University Research School, Ruhr University Bochum, Bochum, Germany
4 DFG Graduate School 736 "Development and Plasticity of the Nervous System: Molecular, synaptic and cellular mechanisms", Ruhr University Bochum, Bochum, Germany
BMC Neuroscience 2010, 11:133 doi:10.1186/1471-2202-11-133Published: 19 October 2010
Members of the complex N-methyl-D-aspartate receptor (NMDAR) subfamily of ionotropic glutamate receptors (iGluRs) conventionally assemble from NR1 and NR2 subunits, the composition of which determines receptor properties. Hallmark features of conventional NMDARs include the requirement for a coagonist, voltage-dependent block by Mg2+, and high permeability for Ca2+. Both Mg2+ sensitivity and Ca2+ permeability are critically dependent on the amino acids at the N and N+1 positions of NR1 and NR2. The recently discovered NR3 subunits feature an unprecedented glycine-arginine combination at those critical sites within the pore. Diheteromers assembled from NR1 and NR3 are not blocked by Mg2+ and are not permeable for Ca2+.
Employing site-directed mutagenesis of receptor subunits, electrophysiological characterization of mutants in a heterologous expression system, and molecular modeling of the NMDAR pore region, we have investigated the contribution of the unusual NR3 N and N+1 site residues to the unique functional characteristics of receptors containing these subunits. Contrary to previous studies, we provide evidence that both the NR3 N and N+1 site amino acids are critically involved in mediating the unique pore properties. Ca2+ permeability could be rescued by mutating the NR3 N site glycine to the NR1-like asparagine. Voltage-dependent Mg2+ block could be established by providing an Mg2+ coordination site at either the NR3 N or N+1 positions. Conversely, "conventional" receptors assembled from NR1 and NR2 could be made Mg2+ insensitive and Ca2+ impermeable by equipping either subunit with the NR3-like glycine at their N positions, with a stronger contribution of the NR1 subunit.
This study sheds light on the structure-function relationship of the least characterized member of the NMDAR subfamily. Contrary to previous reports, we provide evidence for a critical functional involvement of the NR3 N and N+1 site amino acids, and propose them to be the essential determinants for the unique pore properties mediated by this subunit.