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Physiological role of microbodies in the yeast Trichosporon cutaneum during growth on ethylamine as the source of energy, carbon and nitrogen

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Abstract

Compartmentation of the metabolism of ethylamine in Trichosporon cutaneum X4 was studied in cells, grown on this compound as the sole source of energy, carbon, and nitrogen. Transfer experiments indicated that an amine oxidase is involved in the early metabolism of ethylamine. The synthesis of this enzyme was induced by primary amines and was subject to partial carbon catabolite repression. Repression by ammonium ions was not observed. Adaptation of glucose-grown cells to growth on ethylamine was associated with the development of many microbodies, which developed from already existing organelles present in the inoculum cells and multiplied by division. Cytochemical experiments indicated that the organelles contained amine oxidase and catalase. Therefore, they were considered to play a key role in the metabolism of ethylamine. The physiological significance of the microbodies was investigated by fractionation studies of homogenized protoplasts from ethylamine-grown cells by differential- and sucrose-gradient centrifugation of subcellular organelles. Intact microbodies were only obtained when the isolation procedure was performed at pH 5.8 in the absence of Mg2+-ions. Analysis of the different fractions indicated that the key enzymes of the glyoxylate cycle, namely isocitrate lyase and malate synthase, cosedimented together with catalase and amine oxidase. In addition, activities of malate dehydrogenase, glutamate:oxaloacetate aminotransferase (GOT) and (NAD-dependent) glutamate dehydrogenase were detected in these fractions. Electron microscopy revealed that they mainly contained microbodies. Cytochemical experiments indicated that the above enzymes were all present in the same organelle. These findings suggest that microbodies of ethylamine-grown T. cutaneum X4 produce aspartate, so allowing NADH generated in the oxidation of malate by malate dehydrogenase to be quantitatively reoxidized inside the organelles in a series of reactions involving GOT and glutamate dehydrogenase. Aspartase and fumarase were not detected in the microbodies; activities of these two enzymes were present in the cytoplasm.

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Abbreviations

ABTS:

2,2′-Azino-di(3-ethylbenzthiazoline sulfonate [6])

DTT:

dithiothreitol

GOT:

glutamate:oxaloacetate aminotransferase

DTNB:

5,5-dithiobis-2-nitrobenzoate

DAB:

diaminobenzidine

BSPT:

2-(2-benzothiazolyl)-3-(4-phthalhydrazidyl)-t-styryl-sH-tetrazolium chloride

PF:

convex fracture face

EF:

concave fracture face

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Authors and Affiliations

  1. Laboratory for Electron Microscopy, Biological Centre, University of Groningen, Kerklaan 30, NL-9751 NN, Haren, The Netherlands

    M. Veenhuis

  2. Department of Microbiology, Biological Centre, University of Groningen, Kerklaan 30, NL-9751 NN, Haren, The Netherlands

    I. J. van der Klei & W. Harder

Authors
  1. M. Veenhuis
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  2. I. J. van der Klei
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  3. W. Harder
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Veenhuis, M., van der Klei, I.J. & Harder, W. Physiological role of microbodies in the yeast Trichosporon cutaneum during growth on ethylamine as the source of energy, carbon and nitrogen. Arch. Microbiol. 145, 39–50 (1986). https://doi.org/10.1007/BF00413025

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