Neurochemical Research

, Volume 10, Issue 1, pp 99–106 | Cite as

Association of [14C]glutamate with the protein components of synaptic membrane preparations

  • Leena Turpeenoja
  • Pekka Lähdesmäki
Original Articles
  • 9 Downloads

Abstract

Calf brain synaptic plasma membranes were saturated under extracellular conditions with [14C]glutamic acid and the resulting labelled membranes fractionated with 0.9% NaCl, distilled water,n-butanol-water, 0.05 mol/L NaOH and 0.5% Triton X-100 solutions in this order. Triton X-100 was the most effective solubilizer, liberating altogether about 24% of the membrane proteins, but only 4–7% of the label, while NaOH was the most potent solubilizer for the protein-bound label (50–70%). Slab gel electrophoresis showed significant qualitative differences in the banding patterns of the protein extracts, but only two fractions, a low-molecular weight (around 15 kd) and a high-molecular weight (>200 kd) fraction bound [14C]glutamate.

Keywords

Glutamate Glutamic Acid Banding Pattern Qualitative Difference Protein Component 

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References

  1. 1.
    Michaelis, E. K., Michaelis, M. L., andBoyarsky, L. L. 1974. High-affinity glutamic acid binding to brain synaptic membranes. Biochim. Biophys. Acta 367:338–348.PubMedGoogle Scholar
  2. 2.
    Michaelis, E. K., Michaelis, M. L., Stormann, T. M., Chittenden, W. L., andGrubbs, R. D. 1983. Purification and molecular characterization of the brain synaptic membrane glutamate-binding protein. J. Neurochem. 40:1742–1753.PubMedGoogle Scholar
  3. 3.
    Michaelis, E. K., Chittenden, W. L., Johnson, B. E., Galton, N., andDecedue, C. 1984. Purification, biochemical characterization, binding activity, and selectivity of a glutamate binding protein from bovine brain. J. Neurochem. 42:397–406.PubMedGoogle Scholar
  4. 4.
    Roberts, P. J. 1974. Glutamate receptors in the rat central nervous system. Nature (Lond.) 252:399–400.Google Scholar
  5. 5.
    Michaelis, E. K. 1975. Partial purification and characterization of a glutamate-binding membrane glycoprotein from rat brain. Biochem. Biophys. Res. Comm. 65:1004–1012.PubMedGoogle Scholar
  6. 6.
    Snyder, S. H. 1975. The glycine synaptic receptor in the mammalian central nervous system. British J. Pharm. 53:473–484.Google Scholar
  7. 7.
    Fiszer De Plazas, S., andDe Robertis, E. 1975. Isolation of hydrophobic proteins binding amino acids: γ-Aminobutyric acid binding in the rat cerebral cortex. J. Neurochem. 25:547–552.PubMedGoogle Scholar
  8. 8.
    Fiszer De Plazas, S., andDe Robertis, E. 1976. Isolation of hydrophobic proteins binding amino acids:l-aspartic acid-binding protein from the rat cerebral cortex. J. Neurochem. 27:889–894.PubMedGoogle Scholar
  9. 9.
    De Robertis, E., andFiszer De Plazas, S. 1976. Isolation of hydrophobic proteins binding amino acids. Stereoselectivity of the binding ofl-[14C]glutamic acid in cerebral cortex. J. Neurochem. 26:1237–1243.PubMedGoogle Scholar
  10. 10.
    Chude, O. 1979. Solubilization and partial purification of the GABA receptor from mouse brain and a binding assay for the solubilized receptor. J. Neurochem. 33:621–629.PubMedGoogle Scholar
  11. 11.
    Gavish, M., Change, R. S. L., andSnyder, S. H. 1979. Solubilization of histamine H-1, GABA and benzodiazepine receptors. Life Sci. 25:783–790.PubMedGoogle Scholar
  12. 12.
    Greenlee, D. V., andOlsen, R. W. 1979. Solubilization of gamma-aminobutyric acid receptor protein from mammalian brain. Biochem. Biophys. Res. Comm. 88:380–387.PubMedGoogle Scholar
  13. 13.
    Kumpulainen, E., Olkinuora, M., andLähdesmäki, P. 1979. Ability of calf brain synaptic membranes to bind [35S]taurine to their Triton X-100 and chloroform extracts. Neurosci. Lett. 11:215–218.PubMedGoogle Scholar
  14. 14.
    Asano, T., andOgasawara, N. 1980. Solubilization of γ-aminobutyric acid receptor from rat brain. Life Sci. 26:1131–1137.PubMedGoogle Scholar
  15. 15.
    Kanner, B. I., andSharon, I. 1978. Active transport ofl-glutamate by membrane vesicles isolated from rat brain. Biochemistry 17:3949–3953.PubMedGoogle Scholar
  16. 16.
    Young, A. B., andSnyder, S. H. 1974. Strychnine binding in rat spinal cord membranes associated with the synaptic glycine receptors. Cooperativity of glycine interactions. Molec. Pharm. 10:790–809.Google Scholar
  17. 17.
    Zukin, S. R., Young, A. B., andSnyder, S. H. 1974. γ-Aminobutyric acid binding to receptor sites in the rat central nervous system. Proc. Natl. Acad. Sci. U.S.A. 71:4802–4807.PubMedGoogle Scholar
  18. 18.
    Enna, S. J., andSnyder, S. H. 1975. Properties of γ-aminobutyric acid (GABA) receptor binding in rat brain synaptic membrane fractions. Brain Res. 100:81–97.PubMedGoogle Scholar
  19. 19.
    Enna, S. J., andSnyder, S. H. 1977. Influence of ions, enzymes and detergents in γ-aminobutyric acid receptor binding in synaptic membranes of rat brain. Molec. Pharm. 13:442–453.PubMedGoogle Scholar
  20. 20.
    Snyder, S. H., andBennett, J. P. 1976. Neurotransmitter receptors in the brain: Biochemical identification. Annu. Rev. Physiol. 38:153–175.PubMedGoogle Scholar
  21. 21.
    Kadota, K., andKadota, T. 1973. Isolation of coated vesicles, plain synaptic vesicles and flocculent material from a crude synaptosome fraction of guinea pig whole brain. J. Cell Biol. 58:135–151.PubMedGoogle Scholar
  22. 22.
    Tanaka, R., Asaga, H., andTakeda, M. 1976. Nucleoside triphosphate and cation requirement for dopamine uptake by plain synaptic vesicles isolated from rat cerebrums. Brain Res. 115:273–283.PubMedGoogle Scholar
  23. 23.
    Tsudzuki, T. 1979. Mg2+-dependent adenosine triphosphatase in isolated synaptic vesicles. Attribution of most of the enzymic activity to an actomyosin-like protein. J. Biochem. 85:567–574.PubMedGoogle Scholar
  24. 24.
    Turpeenoja, L., andLähdesmäki, P. 1983. Presynaptic binding of amino acids: Characterization of the binding and dissociation properties of taurine, GABA, glutamate, tyrosine and norleucine. Int. J. Neurosci. 22:99–106.PubMedGoogle Scholar
  25. 25.
    Lowry, O. H., Rosebrough, N. J., Farr, A. L., andRandall, R. J. 1951. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193:265–275.PubMedGoogle Scholar
  26. 26.
    Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature (Lond.) 227:680–685.Google Scholar
  27. 27.
    Ratner, N., andMahler, H. 1983. Isolation of postsynaptic densities retaining their membrane attachment. Neurosci. 9:631–644.Google Scholar
  28. 28.
    Lähdesmäki, P., Pasula, M., andOja, S. S. 1975. Effect of electrical stimulation and chlorpromazine on the uptake and release of taurine, γ-aminobutyric acid and glutamic acid in mouse brain synaptosomes. J. Neurochem. 25:675–680.PubMedGoogle Scholar
  29. 29.
    Lähdesmäki, P., Kumpulainen, E., Raasakka, O., andKyrki, P. 1977. Interaction of taurine, GABA and glutamic acid with synaptic membranes. J. Neurochem. 29:819–826.PubMedGoogle Scholar
  30. 30.
    Karppinen, A., Kumpulainen, E., andLähdesmäki, P. 1979. Comparative studies on the interaction of amino acids with synaptosomes, synaptic vesicles and synaptic membranes. Cell. Molec. Biol. 25:335–343.Google Scholar
  31. 31.
    Kumpulainen, E., Karppinen, A., Marnela, K.-M., andLähdesmäki, P. 1981. Subcellular distribution of intracerebrally injected taurine and certain other amino acids in mouse brain. Pages 135–143,in Schaffer, S. W., Baskin, S. I., andKocsis, J. J. (eds.). The Effects of Taurine on Excitable Tissues. Spectrum Publications. Wexford Terrace, Jamaica, New York.Google Scholar
  32. 32.
    Smith, A. P., andLoh, H. H. 1979. Architecture of the nerve ending membrane. Life Sci. 24:1–20.PubMedGoogle Scholar

Copyright information

© Plenum Publishing Corporation 1985

Authors and Affiliations

  • Leena Turpeenoja
    • 1
  • Pekka Lähdesmäki
    • 1
  1. 1.Department of BiochemistryUniversity of OuluOulu 57Finland

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