1887

Abstract

A novel assay method for -aconitate decarboxylase (EC 4.1.1.6) has been developed. Differential centrifugation of cell-free extracts of prepared by Novozyme treatment/nitrogen cavitation of itaconate-producing mycelia has shown that -aconitate decarboxylase, a key enzyme of itaconate biosynthesis, is localized exclusively in the cytosol. The intracellular localization and maximal activities of nine other enzymes putatively involved in itaconate biosynthesis were examined under both producing and non-producing growth conditions. Two striking differences between growth and production mycelia were observed: () -aconitate decarboxylase was absent from growing mycelia, and () aconitase activity was increased at least two-fold in production mycelia. No significant variations in the maximal activities or intracellular localization were observed for the other enzymes investigated. Mitochondrial and cytosolic isoenzymes of NADP:isocitrate dehydrogenase and malate dehydrogenase were detected by cellulose acetate strip electrophoretic analysis, whereas this analysis detected only single isoenzymes of aconitase and citrate synthase. We therefore propose that in itaconate accumulation -aconitate is synthesized in the mitochondrion and then transported to the cytosol where it is converted to itaconate by -aconitate decarboxylase.

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/content/journal/micro/10.1099/00221287-137-3-533
1991-03-01
2021-10-16
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References

  1. Bentley R., Thiessen C. P. 1957a; Biosynthesis of itaconic acid in Aspergillus terreus. I. Tracer studies with 14C-labelled substrates. Journal of Biological Chemistry 226:673–687
    [Google Scholar]
  2. Bentley R., Thiessen C. P. 1957b; Biosynthesis of itaconic acid in Aspergillus terreus. II. Early stages in glucose dissimilation and the role of citrate. Journal of Biological Chemistry 226:689–701
    [Google Scholar]
  3. Bentley R., Thiessen C. P. 1957c; Biosynthesis of itaconic acid in Aspergillus terreus. III. The properties and reaction mechanism of cis-aconitic decarboxylase. Journal of Biological Chemistry 226:703–720
    [Google Scholar]
  4. Bercovitz A., Peleg Y., Battat E., Rokem J. S., Goldberg I. 1990; Localization of pyruvate carboxylase in organic acid-producing Aspergillus strains. Applied and Environmental Microbiology 56:1594–1597
    [Google Scholar]
  5. Bradford M. M. 1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 72:248–254
    [Google Scholar]
  6. De Duve C, Pressman B. C, Gianetto R., Wattiaux R., Appelmans F. 1955; Tissue fractionation studies. 6. Intracellular distribution patterns of enzymes in rat-liver tissue. Biochemical Journal 60:604–617
    [Google Scholar]
  7. Eimhjellen K. E., Larsen H. 1955; The mechanism of itaconic acid formation by Aspergillus terreus. II. The effect of substrates and inhibitors. Biochemical Journal 60:139–147
    [Google Scholar]
  8. Kubicek C. P. 1988; The role of the citric acid cycle in fungal organic acid fermentations. Biochemical Society Symposia 54:113–126
    [Google Scholar]
  9. Kubicek C. P., Róhr M. 1978; The role of the tricarboxylic acid cycle in citric acid accumulation by Aspergillus niger . European Journal of Applied Microbiology and Biotechnology 5:263–271
    [Google Scholar]
  10. Kubicek C. P., Schreferl-Kunar G., Wòhrer W., Rohr M. 1988; Evidence for a cytoplasmic pathway of oxalate biosynthesis in Aspergillus niger . Applied and Environmental Microbiology 54:633–637
    [Google Scholar]
  11. Lal M., Bhargava P. M. 1962; Reversal by pyruvate of fluoride inhibition in Aspergillus terreus . Biochimica et Biophysica Acta 58:628–630
    [Google Scholar]
  12. Lockwood L. B. 1975; Organic acid production. The Filamentous Fungi I151–157 Smith J. E. Berry D. R. London: Edward Arnold;
    [Google Scholar]
  13. Nowakowska-Waszczuk A. 1973; Utilization of some tricarboxylic acid cycle intermediates by mitochondria and growing mycelium of Aspergillus terreus . Journal of General Microbiology 79:19–29
    [Google Scholar]
  14. Osmani S. A., Scrutton M. C. 1983; The sub-cellular localisation of pyruvate carboxylase and of some other enzymes in Aspergillus nidulans . European Journal of Biochemistry 133:551–560
    [Google Scholar]
  15. Osmani S. A., Scrutton M. C. 1985; The sub-cellular localisation and regulatory properties of pyruvate carboxylase from Rhizopus arrhizus . European Journal of Biochemistry 147:119–128
    [Google Scholar]
  16. Osmani S. A., Marston F. A. O., Selmes I. P., Chapman A. G., Scrutton M. C. 1981; Pyruvate carboxylase from Aspergillus nidulans. Regulatory properties. European Journal of Biochemistry 118:271–278
    [Google Scholar]
  17. Peleg Y., Stieglitz B., Goldberg I. 1988; Malic acid accumulation by Aspergillus flavus. I. Biochemical aspects of acid biosynthesis. Applied Microbiology and Biotechnology 28:69–75
    [Google Scholar]
  18. Purohit H. J., Ratledge C. 1988; Mitochondrial location of pyruvate carboxylase in Aspergillus niger . FEMS Microbiology Letters 55:129–132
    [Google Scholar]
  19. Shimi I. R., Nour el Dein M. S. 1962; Biosynthesis of itaconic acid by Aspergillus terreus . Archivflir Mikrobiologie 44:181–188
    [Google Scholar]
  20. Thomson J. F., Nance S. L., Bush K. J., Szczepanik P. A. 1966; Isotope and solvent effects of deuterium on aconitase. Archives of Biochemistry and Biophysics 117:65–74
    [Google Scholar]
  21. Winskill N. 1983; Tricarboxylic acid cycle activity in relation to itaconic acid biosynthesis by Aspergillus terreus . Journal of General Microbiology 129:2877–2883
    [Google Scholar]
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