Advertisement

Neurochemical Research

, Volume 11, Issue 5, pp 733–744 | Cite as

The subcellular localization of glutamate dehydrogenase (GDH): is GDH a marker for mitochondria in brain?

  • James C. K. Lai
  • Kwan-Fu Rex Sheu
  • Young Tai Kim
  • Donald D. Clarke
  • John P. Blass
Original Articles

Abstract

Glutamate dehydrogenase (GDH, EC 1.4.1.2) has long been used as a marker for mitochondria in brain and other tissues, despite reports indicating that GDH is also present in nuclei of liver and dorsal root ganglia. To examine whether GDH can be used as a marker to differentiate between mitochondria and nuclei in the brain, we have measured GDH by enzymatic activity and on immunoblots in rat brain mitochondria and nuclei which were highly enriched by density-gradient centrifugation methods. The activity of GDH was enriched in the nuclear fraction as well as in the mitochondrial faction, while the activities of other “mitochondrial” enzymes (fumarase, NAD-isocitrate dehydrogenase and pyruvate dehydrogenase complex) were enriched only in the mitochondrial fraction. Immunoblots using polyclonal antibodies against bovine liver GDH confirmed the presence of GDH in the rat brain nuclear and mitochondrial fractions. The GDH in these two subcellular fractions had a very similar molecular weight of 56,000 daltons. The mitochondrial and nuclear GDH differed, however, in their susceptibility to solubilization by detergents and salts. The mitochondrial GDH could be solubilized by extraction with low concentrations of detergents (0.1% Triton X-100 and 0.1% Lubrol PX), while the nuclear GDH could be solubizeded only by elevated concentrations of detergents (0.3% each) plus KCl (>150mM). Our results indicate that GDH is present in both nuclei and mitochondria in rat brain. The notion that GDH may serve as a marker for mitochondria needs to be re-evaluated.

Keywords

Dorsal Root Ganglion Subcellular Fraction Pyruvate Dehydrogenase Glutamate Dehydrogenase Mitochondrial Fraction 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Hogeboom, G. H., andSchneider, W. C. (1953. Intracellular distribution of enzymes. XI. Glutamic dehydrogenase. J. Biol. Chem. 204: 233–238.PubMedGoogle Scholar
  2. 2.
    Di Prisco, G., andGarofano, F. (1975). Crystallization and partial characterization of glutamate dehydrogenase from ox nuclei. Biochemistry 14: 4673–4679.PubMedGoogle Scholar
  3. 3.
    di Prisco, G., andCasola, L. (1975). Detection of structural difference between nuclear and mitochondrial glutamate dehydrogenases by the use of immunoabsorbents. Biochemistry 14: 4679–4683.PubMedGoogle Scholar
  4. 4.
    Kato, G., andLowry, O. H. 1973. Distribution of enzymes between nucleus and cytoplasm of single nerve cell bodies. J. Biol. Chem. 248: 2044–2048.PubMedGoogle Scholar
  5. 5.
    Leong, S. F., andClark, J. B. 1984. Regional development of glutamate dehydrogenase in the rat brain. J. Neurochem. 43: 106–111.PubMedGoogle Scholar
  6. 6.
    Filla, A., De Michele, G., Brescia Morra, V., Palma, V., andCampanella, G. 1984. Clinical and biochemical studies on Friedreich's ataxia. Ital. J. Neurol. Sci. 2(Suppl. 4): 47–54.Google Scholar
  7. 7.
    Berl, S., andClarke, D. D. 1984. The metabolic compartmentation concept. Pages 205–218,in Hertz, L., Kvamme, E., McGeer, E. G., andSchousboe, A. (eds.), Glutamine, glutamate and GABA in the central nervous system, Alan R. Liss, New York.Google Scholar
  8. 8.
    Nicklas, W. J., Nunez, R., Berl, S., andDuvoisin, R. 1979. Neuronal-glial contributions to transmitter amino acid metabolism: Studies with kainic acid-induced lesions of rat striatum. J. Neurochem. 33: 839–844.PubMedGoogle Scholar
  9. 9.
    Dennis, S. C., Lai, J. C. K., andClark, J. B. 1977. Comparative studies of glutamale metabolism in synaptic and nonsynaptic rat brain mitochondria. Biochem. J. 164: 727–736.PubMedGoogle Scholar
  10. 10.
    Lai, J. C. K., Sheu, K.-F. R., Clarke, D. D., Kim, Y. T., andBlass, J. P. 1985. Glutamate dehydrogenase in brain mitochondria and nuclei. Trans. Am. Soc. Neurochem. 16: 136.Google Scholar
  11. 11.
    Lai, J. C. K., Sheu, K.-F. R., Clarke, D. D., Kim, Y. T., andBlass, J. P. 1985. Is glutamate dehydrogenase a mitochondrial marker in brain? J. Neurochem. 44(Suppl.) S158.Google Scholar
  12. 12.
    Lai, J. C. K., andClark, J. B. 1979. Preparation of synaptic and nonsynaptic mitochondria from mammalian brain. Methods Enzymol. 55 (Part F) 51–60.PubMedGoogle Scholar
  13. 13.
    Lai, J. C. K., Sheu, K.-F. R., andCarlson Jr., K. C. 1985. Differences in some of the metabolic properties of mitochondria isolated from cerebral cortex and olfactory bulb of the rat. Brain Res. 343: 52–59.PubMedGoogle Scholar
  14. 14.
    Lai, J. C. K., andClark, J. B. 1976. Preparation and properties of mitochondria derived from synaptosomes. Biochem. J. 154: 423–432.PubMedGoogle Scholar
  15. 15.
    Sheu, K.-F. R., Lai, J. C. K., andBlass, J. P. 1984. Properties and regional distribution of pyruvate dehydrogenase kinase in rat brain. J. Neurochem. 42: 230–236.PubMedGoogle Scholar
  16. 16.
    Weber, K., andHiborn, M. 1975. Protein and sodium dodecyl sulfate: Molecular weight determination on polyacrylamide gels and related procedures. Pages 179–223,in Neurath, H., andHill, R. L. (eds.), The proteins, 3rd edition, Academic Press, New York.Google Scholar
  17. 17.
    Sheu, K.-F. R., andKim, Y. T. 1984. Studies on the bovine brain pyruvate dehydrogenase complex using the antibodies against kidney enzyme complex. J. Neurochem. 43: 1352–1358.PubMedGoogle Scholar
  18. 18.
    Sheu, K.-F. R., Lai, J. C. K., Kim, Y. T., Dorante, G., andBagg, J. 1985. Immunochemical characterization of pyruvate dehydrogenase complex in rat brain. J. Neurochem. 44: 593–599.PubMedGoogle Scholar
  19. 19.
    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
  20. 20.
    Løvtrup-Rein, H., andMcEwen, B. S. 1966. Isolation and fractionation of rat brain nuclei. J. Cell Biol. 30: 405–415.PubMedGoogle Scholar
  21. 21.
    Thompson, R. J. 1973. Studies on RNA synthesis in two populations of nuclei in the mammalian cerebral cortex. J. Neurochem. 21: 19–41.PubMedGoogle Scholar
  22. 22.
    Austoker, J., Cox, D., andMathias, A. P. 1972. Fractionation of nuclei from brain by zonal centrifugation and a study of the ribonucleic acid polymerase activity in the various classes of nuclei. Biochem. J. 129: 1139–1155.PubMedGoogle Scholar
  23. 23.
    Chee, P. Y., Dahl, J. L., andFahien, L. A. 1979. The purification and properties of rat brain glutamate dehydrogenase. J. Neurochem. 33: 53–60.PubMedGoogle Scholar
  24. 24.
    King, K. S., andFrieden, C. 1970. The purification and physicaL properties of glutamate dehydrogenase from rat liver. J. Biol. Chem. 245: 4391–4396.PubMedGoogle Scholar
  25. 25.
    Herzfeld, A., Federman, M., andGreengard, O. 1973. Subcellular morphometric and biochemical analysis of developing rat hepatocytes. J. Cell Biol. 57: 475–483.PubMedGoogle Scholar
  26. 26.
    Franke, W. W., Deumling, B., Ermen, B., Jarash, E. D., andKleinig, H. 1970. Nuclear membranes from mammalian liver. I. Isolation procedure and general characterization. J. Cell. Biol. 46: 379–395.PubMedGoogle Scholar
  27. 27.
    di Prisco, G., andStrecker, H. J. 1970. Glutamate dehydrogenase of nuclear and extranuclear compartments of Chang's liver cells. Eur. J. Biochem. 12: 483–489.PubMedGoogle Scholar
  28. 28.
    Plaitakis, A., Nicklas, W. J., andDesnick, R. J. 1980. Glutamate dehydrogenase deficiency in three patients with spinocerebellar syndrome. Ann. Neurol. 7: 297–303.PubMedGoogle Scholar
  29. 29.
    Duvoisin, R. C., Chokroverty, S., Lepore, F., andNicklas, W. 1983. Glutamate dehydrogenase deficiency in patients with olivopontocerebellar atrophy. Neurology (Cleveland) 33: 1322–1326.Google Scholar
  30. 30.
    Plaitakis, A., Berl, S., andYahr, M. D. 1984. Neurological disorders associated with deficiency of glutamate dehydrogenase. Ann. Neurol. 15: 144–153.PubMedGoogle Scholar
  31. 31.
    Sheu, K.-F. R., Cedarbaum, J., Harding, B. J., Clarke, D. D., andBlass, J. P. 1985. Measurements of pyruvate and glutamate dehydrogenases as well fu fumarase in platelet lysates from patients with Parkinson's and related diseases. VIII. Intl. Sym. Parkinson's Disease (Abstract): p38.Google Scholar

Copyright information

© Plenum Publishing Corporation 1986

Authors and Affiliations

  • James C. K. Lai
    • 1
    • 2
  • Kwan-Fu Rex Sheu
    • 1
    • 2
  • Young Tai Kim
    • 3
  • Donald D. Clarke
    • 1
    • 2
  • John P. Blass
    • 1
    • 2
  1. 1.Department of NeurologyCornell University Medical CollegeNew York
  2. 2.Altschul Laboratory for Dementia ResearchBurke Rehabilitation CenterWhite Plains
  3. 3.Department of MedicineCornell University Medical CollegeNew York

Personalised recommendations