Hostname: page-component-cd9895bd7-hc48f Total loading time: 0 Render date: 2024-12-25T03:23:13.350Z Has data issue: false hasContentIssue false
  • English
  • Français

Purification and partial characterization of malate dehydrogenase (decarboxylating) from Tritrichomonas foetus hydrogenosomes

Published online by Cambridge University Press:  06 April 2009

I. hrdý  and
E. Mertens
I. hrdý
Affiliation:
Department of Parasitology, Charles University, Prague, Czech Republic
E. Mertens
Affiliation:
Laboratoire de Chimie Physiologique, Universite Catholique de Louvain and International Institute of Cellular and Molecular Pathology, Brussels, Belgium
Get access Rights & Permissions [Opens in a new window]

Summary

Malate dehydrogenase (decarboxylating) from Tritrichomonas foetus hydrogenosomes was purified close to homogeneity by a combination of differential centrifugation, zwitterionic detergent solubilization, Red-Sepharose chromatography and anion-exchange chromatography. The enzyme with apparent subunit size of 59 kDa and native molecular mass of 308 kDa utilized NAD+ preferentially to NADP+ as a cofactor and required Mn2+ or Mg2+ for its activity. Affinity curves for malate and coenzymes were hyperbolic. Km for malate was 100 μM and 458 μM in the presence of NAD+ and NADP+, respectively. Km for NAD+ and for NADP+ in the presence of malate was 18 μM and 207 μM, respectively. The enzyme is proposed to be a tetramer with a possible physiological role in the maintenance of an appropriate NAD+/NADH ratio in hydrogenosomes.

Keywords

Tritrichomonas foetustrichomonadshydrogenosomesmalic enzyme
Type
Research Article
Information
Parasitology , Volume 107 , Issue 4 , November 1993 , pp. 379 - 385
Copyright
Copyright © Cambridge University Press 1993

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Bradford, M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of proteins utilizing the principle of protein-dye binding. Analytical Biochemistry 77, 248–54.CrossRefGoogle Scholar
Čerkasov, J., Čerkasovová, A. & Dohnalová, M. (1977). Malate dehydrogenases in the cell of Tritrichomonas foetus. Journal of Protozoology 24, 49 A (abstract).Google Scholar
Čerkasov, J., Čerkasovová, A., Kulda, J. & Vilhelmová;, D. (1978). Respiration of hydrogenosomes of Tritrichomonas foetus. I. ADP-dependent oxidation of malate and pyruvate. Journal of Biological Chemistry 253, 1207–14.CrossRefGoogle ScholarPubMed
Čerkasovová, A., Čerkasov, J. & Kulda, J. (1984). Metabolic differences between metronidazole resistant and susceptible strains of Tritrichomonas foetus. Molecular and Biochemical Parasitology 11, 105–18.CrossRefGoogle ScholarPubMed
Diamond, L. S. (1957). The establishment of various trichomonads of animals and man in axenic cultures. Journal of Parasitology 43, 488–90.CrossRefGoogle ScholarPubMed
Fodge, D. W., Gracy, R. W. & Harris, B. G. (1972). Studies on enzymes from parasitic helminths. I. Purification and physical properties of malic enzyme from muscle tissue of Ascaris suum. Biochimica et Biophysica Acta 268, 271–84.CrossRefGoogle ScholarPubMed
Honigberg, B. M. (1978 a). Trichomonads of importance in human medicine. In Parasitic Protozoa, vol. 2 (ed. Kreier, J. P.), pp. 275–454. New York: Academic Press.Google Scholar
Honigberg, B. M. (1978 b). Trichomonads of veterinary importance. In Parasitic Protozoa, vol. 2 (ed. Kreier, J. P.), pp. 163–273. New York: Academic Press.Google Scholar
Hrdý, I., Mertens, E. & Van Schaftingen, E. (1993). Identification, purification and separation of different isozymes of NADP-specific malic enzyme from Tritrichomonas foetus. Molecular and Biochemical Parasitology 57, 253–60.CrossRefGoogle ScholarPubMed
Kulda, J., Nohýnková, E. & Ludvík, J. (1987). Basic structure of the trichomonad cell. Acta Universitatis Carolinae – Biologica 30, 181–98.Google Scholar
Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, London 227, 680–5.CrossRefGoogle ScholarPubMed
Lindmark, D. G., Dinbergs, I. D., Halberg, L. A. & Eckenrode, B. L. (1989 a). Purification of hydrogenosomal enzymes from Tritrichomonas foetus. In Biochemistry and Molecular Biology of ‘Anaerobic’ Protozoa (ed. Lloyd, D., Coombs, G. H. & Paget, T. A.), pp. 161–71. London: Harwood Academic Publishers.Google Scholar
Lindmark, D. G., Eckenrode, B. L., Halberg, L. A. & Dinbergs, I. D. (1989 b). Carbohydrate, energy and hydrogenosomal metabolism of Tritrichomonas foetus and Trichomonas vaginalis. Journal of Protozoology 36, 214–16.CrossRefGoogle ScholarPubMed
Lindmark, D. G. & Müller, M. (1973). Hydrogenosome, a cytoplasmic organelle of the anaerobic flagellate, Tritrichomonas foetus, and its role in pyruvate metabolism. Journal of Biological Chemistry 248, 7724–8.CrossRefGoogle ScholarPubMed
Müller, M. (1973). Biochemical cytology of trichomonas flagellates. I. Subcellular localization of hydrolases, dehydrogenases, and catalase in Tritrichomonas foetus. Journal of Cell Biology 57, 453–74.Google Scholar
Müller, M. (1976). Carbohydrate and energy metabolism of Tritrichomonas foetus. In Biochemistry of Parasites and Host–Parasite Relationships (ed. Van den Bossche, H.), pp. 3–14. Amsterdam: North-Holland Publishing company.Google Scholar
Müller, M. (1980). The hydrogenosome. In The Eukaryotic Microbial Cell (ed. Gooday, G. W., Lloyd, D. & Trinci, A. P. J.). Society for General Microbiology Symposium 30, pp. 127–147. Cambridge: Cambridge University Press.Google Scholar
Müller, M. (1988). Energy metabolism of protozoa without mitochondria. Annual Review of Microbiology 42, 465–88.CrossRefGoogle ScholarPubMed
Nyren, P., Nore, B. F. & Strid, A. (1991). Proton-pumping N, N'-dicyclohexylcarbodiimide-sensitive inorganic pyrophosphate synthase from Rhodospirillum rubrum. Purification, characterization and reconstitution. Biochemistry 30, 2883–7.CrossRefGoogle ScholarPubMed
Sauer, L. A. (1973). Mitochondrial NAD-dependent malic enzyme: a new regulatory enzyme. FEBS Letters 33, 251–5.CrossRefGoogle ScholarPubMed
Steinbüchel, A. & Müller, M. (1986 a). Glycerol, a metabolic end product of Trichomonas vaginalis and Tritrichomonas foetus. Molecular and Biochemical Parasitology 20, 45–55.CrossRefGoogle ScholarPubMed
Steinbüchel, A. & Müller, M. (1986 b). Anaerobic pyruvate metabolism of Tritrichomonas foetus and Trichomonas vaginalis hydrogenosomes. Molecular and Biochemical Parasitology 20, 57–65.CrossRefGoogle ScholarPubMed
Tanabe, M., Miura, S., Kaneda, Y., Takeuchi, T. & Asami, K. (1980). Trichomonas vaginalis: Subcellular localization of NAD+- and NADP+-linked malic enzymes. Japanese Journal of Parasitology 29, 19–26.Google Scholar
Taroni, F. & Di-Donato, S. (1988). Purification and properties of cytosolic malic enzyme from human skeletal muscle. International Journal of Biochemistry 20, 857–66.CrossRefGoogle ScholarPubMed