Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1996 Aug 20;93(17):9259–9263. doi: 10.1073/pnas.93.17.9259

A mutation that alters magnesium block of N-methyl-D-aspartate receptor channels.

G Sharma 1, C F Stevens 1
PMCID: PMC38629  PMID: 8799188

Abstract

N-Methyl-D-aspartate (NMDA) receptors are blocked at hyperpolarizing potentials by extracellular Mg ions. Here we present a detailed kinetic analysis of the Mg block in recombinant wild-type and mutant NMDA receptors. We find that the Mg binding site is the same in the wild-type and native hippocampal NMDA receptor channels. In the mutant channels, however, Mg ions bind with a 10-fold lower affinity. On the basis of these results, we propose that the energy well at the Mg binding site in the mutants is shallow and the binding is unstable because of an increase in the rate of dissociation. We postulate that the dipole formed by the amide group of asparagine 614 of the epsilon 1 subunit contributes to the structure of the binding site but predict that additional ligands will be involved in coordinating Mg ions.

Full text

PDF
9259

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ascher P., Nowak L. The role of divalent cations in the N-methyl-D-aspartate responses of mouse central neurones in culture. J Physiol. 1988 May;399:247–266. doi: 10.1113/jphysiol.1988.sp017078. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bekkers J. M., Stevens C. F. NMDA receptors at excitatory synapses in the hippocampus: test of a theory of magnesium block. Neurosci Lett. 1993 Jun 25;156(1-2):73–77. doi: 10.1016/0304-3940(93)90443-o. [DOI] [PubMed] [Google Scholar]
  3. Burnashev N., Schoepfer R., Monyer H., Ruppersberg J. P., Günther W., Seeburg P. H., Sakmann B. Control by asparagine residues of calcium permeability and magnesium blockade in the NMDA receptor. Science. 1992 Sep 4;257(5075):1415–1419. doi: 10.1126/science.1382314. [DOI] [PubMed] [Google Scholar]
  4. Chen C., Okayama H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol. 1987 Aug;7(8):2745–2752. doi: 10.1128/mcb.7.8.2745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Collingridge G. L., Lester R. A. Excitatory amino acid receptors in the vertebrate central nervous system. Pharmacol Rev. 1989 Jun;41(2):143–210. [PubMed] [Google Scholar]
  6. Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F. J. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 1981 Aug;391(2):85–100. doi: 10.1007/BF00656997. [DOI] [PubMed] [Google Scholar]
  7. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. doi: 10.1016/0378-1119(89)90358-2. [DOI] [PubMed] [Google Scholar]
  8. Hollmann M., Maron C., Heinemann S. N-glycosylation site tagging suggests a three transmembrane domain topology for the glutamate receptor GluR1. Neuron. 1994 Dec;13(6):1331–1343. doi: 10.1016/0896-6273(94)90419-7. [DOI] [PubMed] [Google Scholar]
  9. Jahr C. E., Stevens C. F. A quantitative description of NMDA receptor-channel kinetic behavior. J Neurosci. 1990 Jun;10(6):1830–1837. doi: 10.1523/JNEUROSCI.10-06-01830.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jahr C. E., Stevens C. F. Voltage dependence of NMDA-activated macroscopic conductances predicted by single-channel kinetics. J Neurosci. 1990 Sep;10(9):3178–3182. doi: 10.1523/JNEUROSCI.10-09-03178.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kawajiri S., Dingledine R. Multiple structural determinants of voltage-dependent magnesium block in recombinant NMDA receptors. Neuropharmacology. 1993 Nov;32(11):1203–1211. doi: 10.1016/0028-3908(93)90014-t. [DOI] [PubMed] [Google Scholar]
  12. Mayer M. L., Westbrook G. L., Guthrie P. B. Voltage-dependent block by Mg2+ of NMDA responses in spinal cord neurones. Nature. 1984 May 17;309(5965):261–263. doi: 10.1038/309261a0. [DOI] [PubMed] [Google Scholar]
  13. Meguro H., Mori H., Araki K., Kushiya E., Kutsuwada T., Yamazaki M., Kumanishi T., Arakawa M., Sakimura K., Mishina M. Functional characterization of a heteromeric NMDA receptor channel expressed from cloned cDNAs. Nature. 1992 May 7;357(6373):70–74. doi: 10.1038/357070a0. [DOI] [PubMed] [Google Scholar]
  14. Monyer H., Sprengel R., Schoepfer R., Herb A., Higuchi M., Lomeli H., Burnashev N., Sakmann B., Seeburg P. H. Heteromeric NMDA receptors: molecular and functional distinction of subtypes. Science. 1992 May 22;256(5060):1217–1221. doi: 10.1126/science.256.5060.1217. [DOI] [PubMed] [Google Scholar]
  15. Mori H., Masaki H., Yamakura T., Mishina M. Identification by mutagenesis of a Mg(2+)-block site of the NMDA receptor channel. Nature. 1992 Aug 20;358(6388):673–675. doi: 10.1038/358673a0. [DOI] [PubMed] [Google Scholar]
  16. Moriyoshi K., Masu M., Ishii T., Shigemoto R., Mizuno N., Nakanishi S. Molecular cloning and characterization of the rat NMDA receptor. Nature. 1991 Nov 7;354(6348):31–37. doi: 10.1038/354031a0. [DOI] [PubMed] [Google Scholar]
  17. Needham J. V., Chen T. Y., Falke J. J. Novel ion specificity of a carboxylate cluster Mg(II) binding site: strong charge selectivity and weak size selectivity. Biochemistry. 1993 Apr 6;32(13):3363–3367. doi: 10.1021/bi00064a020. [DOI] [PubMed] [Google Scholar]
  18. Nowak L., Bregestovski P., Ascher P., Herbet A., Prochiantz A. Magnesium gates glutamate-activated channels in mouse central neurones. Nature. 1984 Feb 2;307(5950):462–465. doi: 10.1038/307462a0. [DOI] [PubMed] [Google Scholar]
  19. Premkumar L. S., Auerbach A. Identification of a high affinity divalent cation binding site near the entrance of the NMDA receptor channel. Neuron. 1996 Apr;16(4):869–880. doi: 10.1016/s0896-6273(00)80107-5. [DOI] [PubMed] [Google Scholar]
  20. Ruppersberg J. P., Mosbacher J., Günther W., Schoepfer R., Fakler B. Studying block in cloned N-methyl-D-aspartate (NMDA) receptors. Biochem Pharmacol. 1993 Dec 3;46(11):1877–1885. doi: 10.1016/0006-2952(93)90627-9. [DOI] [PubMed] [Google Scholar]
  21. Sakurada K., Masu M., Nakanishi S. Alteration of Ca2+ permeability and sensitivity to Mg2+ and channel blockers by a single amino acid substitution in the N-methyl-D-aspartate receptor. J Biol Chem. 1993 Jan 5;268(1):410–415. [PubMed] [Google Scholar]
  22. Stern P., Béhé P., Schoepfer R., Colquhoun D. Single-channel conductances of NMDA receptors expressed from cloned cDNAs: comparison with native receptors. Proc Biol Sci. 1992 Dec 22;250(1329):271–277. doi: 10.1098/rspb.1992.0159. [DOI] [PubMed] [Google Scholar]
  23. Stern P., Cik M., Colquhoun D., Stephenson F. A. Single channel properties of cloned NMDA receptors in a human cell line: comparison with results from Xenopus oocytes. J Physiol. 1994 May 1;476(3):391–397. doi: 10.1113/jphysiol.1994.sp020140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Tsuzuki K., Mochizuki S., Iino M., Mori H., Mishina M., Ozawa S. Ion permeation properties of the cloned mouse epsilon 2/zeta 1 NMDA receptor channel. Brain Res Mol Brain Res. 1994 Oct;26(1-2):37–46. doi: 10.1016/0169-328x(94)90071-x. [DOI] [PubMed] [Google Scholar]
  25. Villarroel A., Burnashev N., Sakmann B. Dimensions of the narrow portion of a recombinant NMDA receptor channel. Biophys J. 1995 Mar;68(3):866–875. doi: 10.1016/S0006-3495(95)80263-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Yamazaki M., Mori H., Araki K., Mori K. J., Mishina M. Cloning, expression and modulation of a mouse NMDA receptor subunit. FEBS Lett. 1992 Mar 23;300(1):39–45. doi: 10.1016/0014-5793(92)80160-i. [DOI] [PubMed] [Google Scholar]
  27. Zarei M. M., Dani J. A. Structural basis for explaining open-channel blockade of the NMDA receptor. J Neurosci. 1995 Feb;15(2):1446–1454. doi: 10.1523/JNEUROSCI.15-02-01446.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES