Identification of a new site in the S1 ligand binding region of the NMDA receptor NR2A subunit involved in receptor activation by glutamate.
Activation of N-methyl-d-aspartate (NMDA) receptors requires the binding of both glutamate and glycine to independent sites on the receptor. These ligands bind to NR2 and NR1 subunits respectively. Ligand binding residues are located in two non-contiguous domains, S1 and S2, which have been implicated in glutamate binding in other ionotropic glutamate receptor subunits. To further define the amino acids through which glutamate activates the receptor, we generated single-site mutations to the NR2A subunit, and expressed them with wild type NR1 in HEK 293 cells. Using calcium imaging and whole cell patch clamp we determined glutamate and glycine potencies. Of the eight residues mutated we identified five (E413, K484, A508, G685 and G688), whose mutation leads to a large reduction (from 4- to 1000-fold) in glutamate potency, consistent with a role for these residues in receptor activation by glutamate. The potency of glycine was largely unchanged by these mutations. Thus our results extend the knowledge base of residues involved in NMDA receptor function and identifies a new site in S1, in the region of A508, that has a role in receptor activation by glutamate.
Lummis
Sarah C R
SC
Nature Biotechnology, London, UK. sl120@melo.bio.cam.ac.uk
Fletcher
Elizabeth J
EJ
Green
Tim
T
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D018698:Glutamic Acid / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D017354:Point Mutation / Q000502:physiology; D051381:Rats; D016194:Receptors, N-Methyl-D-Aspartate / Q000235:genetics / Q000378:metabolism*
D018698:Glutamic Acid; D008024:Ligands; D016194:Receptors, N-Methyl-D-Aspartate; C409466:NR1 NMDA receptor