Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Jan;161(1):265–271. doi: 10.1128/jb.161.1.265-271.1985

Presence and regulation of the alpha-ketoglutarate dehydrogenase multienzyme complex in the filamentous fungus Aspergillus niger.

B Meixner-Monori, C P Kubicek, A Habison, E M Kubicek-Pranz, M Röhr
PMCID: PMC214866  PMID: 3968029

Abstract

alpha-Ketoglutarate dehydrogenase has been demonstrated for the first time in cell extracts from the filamentous fungus Aspergillus niger. A minimum protein concentration of 5 mg/ml is necessary for detecting enzyme activity, but a maximum of ca. 0.060 mumol/min per mg of protein is observed only when the protein concentration is above 9 mg/ml. alpha-Ketoglutarate can partly stabilize the enzyme against dilution in the assay system. Neither bovine serum albumin nor a variety of substrates or effectors of the enzyme could stabilize the enzyme against inactivation by dilution. A kinetic analysis of the enzyme revealed Michaelis-Menten kinetics with respect to alpha-ketoglutarate, coenzyme A, and NAD. Thiamine PPi was required for maximal activity. NADH, oxaloacetate, succinate, and cis-aconitate were found to inhibit the enzyme; AMP was without effect. Monovalent cations including NH4+ were inhibitory at high concentrations (greater than 20 mM). The highest enzyme activity was found in rapidly growing mycelia (glucose-NH4+ or glucose-peptone medium). We discuss the possibility that citric acid accumulation is caused by oxaloacetate and NADH inhibition of the alpha-ketoglutarate dehydrogenase of A. niger.

Full text

PDF
266

Selected References

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

  1. Amarasingham C. R., Davis B. D. Regulation of alpha-ketoglutarate dehydrogenase formation in Escherichia coli. J Biol Chem. 1965 Sep;240(9):3664–3668. [PubMed] [Google Scholar]
  2. Boonsaeng V., Sullivan P. A., Shepherd M. G. Phosphofrucktokinase and glucose catabolism of Mucor and Penicillium species. Can J Microbiol. 1977 Sep;23(9):1214–1224. doi: 10.1139/m77-182. [DOI] [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Führer L., Kubicek C. P., Röhr M. Pyridine nucleotide levels and ratios in Aspergillus niger. Can J Microbiol. 1980 Mar;26(3):405–408. doi: 10.1139/m80-067. [DOI] [PubMed] [Google Scholar]
  5. Habison A., Kubicek C. P., Röhr M. Partial purification and regulatory properties of phosphofructokinase from Aspergillus niger. Biochem J. 1983 Mar 1;209(3):669–676. doi: 10.1042/bj2090669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hansen H. G., Henning U. Regulation of pyruvate dehydrogenase activity in Escherichia coli K12. Biochim Biophys Acta. 1966 Aug 10;122(2):355–358. doi: 10.1016/0926-6593(66)90076-2. [DOI] [PubMed] [Google Scholar]
  7. Hirashima M., Hayakawa T., Koike M. Mammalian alpha-keto acid dehydrogenase complexes. II. An improved procedure for the preparation of 2-oxoglutarate dehydrogenase complex from pig heart muscle. J Biol Chem. 1967 Mar 10;242(5):902–907. [PubMed] [Google Scholar]
  8. Ishikawa E., Oliver R. M., Reed L. J. Alpha-Keto acid dehydrogenase complexes, V. Macromolecular organization of pyruvate and alpha-ketoglutarate dehydrogenase complexes isolated from beef kidney mitochondria. Proc Natl Acad Sci U S A. 1966 Aug;56(2):534–541. doi: 10.1073/pnas.56.2.534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Khouw B. T., McCurdy H. D. Tricarboxylic acid cycle enzymes and morphogenesis in Blastocladiella emersonii. J Bacteriol. 1969 Jul;99(1):197–205. doi: 10.1128/jb.99.1.197-205.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kornfeld S., Benziman M., Milner Y. Alpha-ketoglutarate dehydrogenase complex of Acetobacter xylinum. Purification and regulatory properties. J Biol Chem. 1977 May 10;252(9):2940–2947. [PubMed] [Google Scholar]
  11. Kubicek C. P., Röhr M. Regulation of citrate synthase from the citric acid-accumulating fungus, Aspergillus niger. Biochim Biophys Acta. 1980 Oct;615(2):449–457. [PubMed] [Google Scholar]
  12. LaNoue K. F., Walajtys E. I., Williamson J. R. Regulation of glutamate metabolism and interactions with the citric acid cycle in rat heart mitochondria. J Biol Chem. 1973 Oct 25;248(20):7171–7183. [PubMed] [Google Scholar]
  13. Meixner-Monori B., Kubicek C. P., Röhr M. Pyruvate kinase from Aspergillus niger: a regulatory enzyme in glycolysis? Can J Microbiol. 1984 Jan;30(1):16–22. doi: 10.1139/m84-003. [DOI] [PubMed] [Google Scholar]
  14. Osmani S. A., Scrutton M. C. The sub-cellular localisation of pyruvate carboxylase and of some other enzymes in Aspergillus nidulans. Eur J Biochem. 1983 Jul 1;133(3):551–560. doi: 10.1111/j.1432-1033.1983.tb07499.x. [DOI] [PubMed] [Google Scholar]
  15. Parker M. G., Weitzman P. D. The purification and regulatory properties of alpha-oxoglutarate dehydrogenase from Acineobacter iwoffi. Biochem J. 1973 Sep;135(1):215–223. doi: 10.1042/bj1350215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Poulsen L. L., Wedding R. T. Purification and properties of the alpha-ketoglutarate dehydrogenase complex of cauliflower mitochondria. J Biol Chem. 1970 Nov 10;245(21):5709–5717. [PubMed] [Google Scholar]
  17. Ryan L. D., Vestling C. S. Rapid purification of lactate dehydrogenase from rat liver and hepatoma: a new approach. Arch Biochem Biophys. 1974 Jan;160(1):279–284. doi: 10.1016/s0003-9861(74)80035-4. [DOI] [PubMed] [Google Scholar]
  18. Smith C. M., Bryla J., Williamson J. R. Regulation of mitochondrial alpha-ketoglutarate metabolism by product inhibition at alpha-ketoglutarate dehydrogenase. J Biol Chem. 1974 Mar 10;249(5):1497–1505. [PubMed] [Google Scholar]
  19. Strumilo S. A., Taranda N. I., Senkevich S. B., Vinogradov V. V. 2-oxoglutarate dehydrogenase complex from bovine-adrenal-cortex mitochondria. Purification and partial characterization. Acta Biol Med Ger. 1981;40(3):257–264. [PubMed] [Google Scholar]
  20. WAGNER R. P., BERGQUIST A., BARBEE T. THE SYNTHESIS IN VITRO OF VALINE AND ISOLEUCINE FROM PYRUVATE AND ALPHA-KETOBUTYRATE IN NEUROSPORA. Biochim Biophys Acta. 1965 May 4;100:444–450. doi: 10.1016/0304-4165(65)90014-0. [DOI] [PubMed] [Google Scholar]
  21. Watson K., Smith J. E. Oxidative phosphorylation and respiratory control in mitochondria from Aspergillus niger. Biochem J. 1967 Aug;104(2):332–339. doi: 10.1042/bj1040332. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Williamson J. R., Scholz R., Browning E. T. Control mechanisms of gluconeogenesis and ketogenesis. II. Interactions between fatty acid oxidation and the citric acid cycle in perfused rat liver. J Biol Chem. 1969 Sep 10;244(17):4617–4627. [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES