title: DRPEER: a motif in the extracellular vestibule conferring high Ca2+ flux rates in NMDA receptor channels.
abstract: The high flux rate of Ca2+ through NMDA receptor (NMDAR) channels is critical for their biological function and may depend on a Ca2+ binding site in the extracellular vestibule. We screened substitutions of hydrophilic residues exposed in the vestibule and identified a cluster of charged residues and a proline, the DRPEER motif, positioned C terminal to M3, that is unique to the NR1 subunit. Charge neutralization or conversion of residues in DRPEER altered fractional Ca2+ currents in a manner consistent with its forming a binding site for Ca2+. Similarly, in a mutant channel in which all of the negative charges are neutralized (ARPAAR), the block by extracellular Ca2+ of single-channel current amplitudes is attenuated. In these same channels, the block by extracellular Mg2+ is unaffected. DRPEER is located extracellularly, and its contribution to Ca2+ influx is distinct from that of the narrow constriction. We conclude that key residues in DRPEER, acting as an external binding site for Ca2+, along with a conserved asparagine in the M3 segment proper, contribute to the high fractional Ca2+ currents in these channels under physiological conditions. Therefore, these domains represent critical molecular determinants of NMDAR function in synaptic physiology.
authors.affiliation: Graduate Program in Neurobiology and Behavior, State University of New York at Stony Brook, Stony Brook, New York 11794-5230, USA.
authors.forename: Junryo
authors.initials: J
authors.lastname: Watanabe
authors.forename: Christine
authors.initials: C
authors.lastname: Beck
authors.forename: Thomas
authors.initials: T
authors.lastname: Kuner
authors.forename: Louis S
authors.initials: LS
authors.lastname: Premkumar
authors.forename: Lonnie P
authors.initials: LP
authors.lastname: Wollmuth
mesh_terms: D020816:Amino Acid Motifs / Q000502:physiology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D005110:Extracellular Space / Q000378:metabolism*; D006801:Humans; D007473:Ion Channels / Q000378:metabolism*; D017136:Ion Transport / Q000502:physiology*; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D016194:Receptors, N-Methyl-D-Aspartate / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D014981:Xenopus
chemical_list: D007473:Ion Channels; C409466:NR1 NMDA receptor; C120997:NR2A NMDA receptor; C432823:NR2C NMDA receptor; D021122:Protein Subunits; D016194:Receptors, N-Methyl-D-Aspartate; D008274:Magnesium; D002118:Calcium