The amino acids L-glutamate and glycine are essential agonists of the excitatory NMDA receptor, a subtype ionotropic glutamate receptor family. native is composed two types homologous membrane-spanning subunits, NR1 NR2. Here, numbers glycine-binding glutamate-binding NR2 subunits in hetero-oligomer were determined by coexpressing wild-type (wt) with low-affinity mutant NR1(Q387K), wt NR2B NR2BE387A, Xenopus oocytes. In both cases, analysis resulting dose-response curves revealed three...

NMDA receptors require both L-glutamate and the coagonist glycine for efficient channel activation. The binding site of these heteromeric receptor proteins is formed by regions NMDAR1 (NR1) subunit that display sequence similarity to bacterial amino acid proteins. Here, we demonstrate glutamate located on homologous NR2B subunit. Mutation residues within N-terminal domain loop region between membrane segments M3 M4 significantly reduced efficacy glutamate, but not glycine, in gating. Some...

Glycine receptors are anchored at inhibitory chemical synapses by a cytoplasmic protein, gephyrin. Molecular cloning revealed the similarity of gephyrin to prokaryotic and invertebrate proteins essential for synthesizing cofactor required activity molybdoenzymes. Gene targeting in mice showed that is both synaptic clustering glycine spinal cord molybdoenzyme nonneural tissues. The mutant phenotype resembled humans with hereditary molybdenum deficiency hyperekplexia (a failure...

The N-methyl-D-aspartate (NMDA) subtype of ionotropic glutamate receptors is a heterooligomeric membrane protein composed homologous subunits. Here, the contribution M3-M4 loop NR1 subunit to binding and co-agonist glycine was investigated by site-directed mutagenesis. Substitution phenylalanine residues at positions 735 or 736 produced 15- 30-fold reduction in apparent affinity without affecting competitive antagonist 7-chlorokynurenic acid; mutation both caused >100-fold decrease...

By homology screening of a rat brain cDNA library we have identified novel alpha subunit variant the inhibitory glycine receptor, 3. The deduced protein exhibits 82-83% amino acid identity to previously characterized and human 1 2 sequences. Upon heterologous expression in Xenopus laevis oocytes, 3 formed functional agonist-gated chloride channels which displayed low affinity for only small responses taurine. Amplification transcripts by polymerase chain reaction revealed high levels spinal...

1. The effect of the divalent cation Zn2+ on inhibitory glycine receptor (GlyR) currents was investigated in rat embryonic spinal cord neurons and Xenopus oocytes expressing recombinant GlyRs. 2. In cultured neurons, potentiated glycine-induced whole-cell about 3-fold when applied extracellularly at concentrations 0.5-10 microM. contrast, higher (> 100 microM) decreased response. 3. A similar biphasic modulation by also found with homo- hetero-oligomeric GlyRs generated oocytes....

The amino acid at position 160 of the ligand-binding subunit, α1, is an important determinant agonist and antagonist binding to glycine receptor. Exchange neighboring residues, phenylalanine 159 tyrosine 161, increased efficacy agonists. Whereas wild-type α1 channels expressed in Xenopus oocytes required 0.7 millimolar β-alanine for a half-maximal response, doubly mutated (F159Y,Y161F) subunit had affinity (which was more potent than glycine) that 110-fold wild type. Also, γ-aminobutyric...

The inhibitory glycine receptor (GlyR) is a ligand‐gated chloride channel protein which displays developmental heterogeneity in the mammalian central nervous system. Here we describe 2 novel cDNA variants of rat GlyR α2 subunit and demonstrate that alternative splicing generates these isoforms. deduced sequences (α2A α2B) exhibit 99% identity with previously characterized human subunit. In situ hybridization revealed expression both α2A und α2B mRNAs prenatal brain, suggesting variant...

The inhibitory glycine receptor (GlyR) in mammalian spinal cord displays pharmacological and molecular heterogeneity of its strychnine binding alpha subunit. Here, cDNAs were isolated which encode a variant (alpha ins 1) the rat GlyR 1 subunit that contains eight additional amino acids putative cytoplasmic domain. Analysis corresponding genomic sequence showed transcripts result from alternative splice acceptor site selection. S1 nuclease protection experiments, Northern blot analysis, RNA...

Several lines of evidence imply changes in inhibitory interneuron connectivity and subsequent alterations oscillatory network activities the pathogenesis Alzheimer's Disease (AD). Recently, we provided for an increased immunoreactivity both postsynaptic scaffold protein gephyrin GABAA receptor γ2-subunit hippocampus young (1 3 months age), APPPS1 mice. These mice represent a well-established model cerebral amyloidosis, which is hallmark human AD. In this study, demonstrate robust increase...

1. The effects of Zn2+ on glycine receptor (GlyR) currents were analysed in Xenopus oocytes and human embryonic kidney cells expressing homomeric wild-type mutant alpha1 subunit GlyRs. 2. Low concentrations (10 microM) extracellular converted the partial agonist taurine into a high-efficacy agonist. Concentration-response analysis showed that EC50 for decreased whereas Hill coefficient increased under these conditions. In contrast, 50-500 microM an value reduced maximal inducible currents....

Homozygotic spasmodic ( spd/spd ) mice suffer from a motor disorder resembling poisoning by the glycine receptor antagonist strychnine. Here, point mutation was identified in α1 subunit gene of mouse which predicts an alanine‐to‐serine exchange at position 52 mature polypeptide. Upon expression Xenopus laevis oocytes, A52S channels displayed reduced responses to glycine, β‐alanine and taurine when compared recombinant receptors. As content spinal cord native molecular weight appeared...

Gephyrin is a scaffold protein essential for the postsynaptic clustering of inhibitory glycine and different subtypes GABA_A receptors. The cellular molecular mechanisms involved in gephyrin-mediated receptor are still not well understood. Here we provide evidence that gephyrin-binding collybistin regulating phosphorylation gephyrin. We demonstrate widely used monoclonal antibody mAb7a phospho-specific allows biochemical analysis gephyrin at Ser-270. In addition, another neighbored...

Abstract Immunohistochemistry and in situ hybridization were used to study the distribution of glycine receptor (GlyR) subunits GlyR-associated protein gephyrin rat retina. Monoclonal antibodies against α β GlyR showed a strong punctate labeling pattern inner plexiform layer. Glycine mRNAs found nuclear layer ganglion cell The 1 subunit mRNA is predominantly present outer half INL on some but not all cells. α2 nearly cells α3–, β-, gephyrin-mRNAs are entire differential expression indicates...