1887

Abstract

SUMMARY: Thiosulphate dehydrogenase (EC 1.8.2.2; thiosulphate:acceptor oxidoreductase) was purified to apparent homogeneity from by a combination of ammonium sulphate precipitation, hydrophobic interaction chromatography, anion-exchange chromatography and gel filtration. The enzyme catalysed the oxidation of thiosulphate (SO ) to tetrathionate (SO ) with potassium ferricyanide as an artificial electron acceptor. The molecular mass of the native enzyme, as determined by gel filtration, was 102 ± 4.2 kDa. The enzyme contained two different subunits with a molecular mass of 24 ± 0.9 and 20 ± 1.0 kDa (SDS-PAGE), respectively. Both subunits contained -type haem with absorption bands at 553, 524 and 416 nm. A 77 K spectrum of purified thiosulphate dehydrogenase revealed that the absorption at 553 nm is due to different haem groups. A cytochrome content of 5.3 mole -type haem per mole of native enzyme was calculated. The pH optimum of the purified enzyme was 3. Apart from ferricyanide, Wurster's blue (the free radical of tetramethyl -phenylenediamine) and horse heart cytochrome could also serve as electron acceptors, though less effectively than ferricyanide. At pH 7.0, the for thiosulphate was 0.54 mM. The could not be determined at the pH optimum due to the chemical reactivity of thiosulphate at low pH values. Sulphite was a potent inhibitor of enzyme activity.

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1993-09-01
2024-03-28
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References

  1. Bartsch R.G. 1971; Cytochromes: bacterial.. Methods in Enzymology 23:344–363
    [Google Scholar]
  2. 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]
  3. Gorenstein M.V., Li J.B., Chapman D. 1993 Abstract no.974 Pittsburgh Conference; Atlanta, USA:
    [Google Scholar]
  4. Laemmli U.K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4.. Nature; London: 227680–685
    [Google Scholar]
  5. Lane D.J., Harrison A.P. JR Stahl D., Pace B., Giovannonj S.J., Olsen G.J., Pace N.R. 1992; Evolutionary relationships among sulfur- and iron-oxidizing eubacteria.. Journal of Bacteriology 174:269–278
    [Google Scholar]
  6. Lu W.-P., Kelly D.P. 1988; Cellular location and partial purification of the thiosulphate-oxidizing enzyme and trithionate hydrolasefrom Thiobacillus tepidarius.. Journal of General Microbiology 134:877–885
    [Google Scholar]
  7. Lyric R.M., Suzuki I. 1970; Enzymes involved in the metabolism of thiosulfate by Thiobacillus thioparus. III. Properties of thiosulfate- oxidizing enzyme and proposed pathway of thiosulfate oxidation.. Canadian Journal of Biochemistry 48:355–363
    [Google Scholar]
  8. Meulenberg R., Pronk J. T., Hazeu W., Bos P., Kuenen J. G. 1992a; Oxidation of reduced sulphur compounds by intact cells of Thiobacillus acidophilus.. Archives of Microbiology 157:161–168
    [Google Scholar]
  9. Meulenberg R., Pronk J. T., Frank J., Hazeu W., Bos P., Kuenen J.G. 1992b; Purification and partial characterization of a thermostable trithionate hydrolase from the acidophilic sulphur oxidizer Thiobacillus acidophilus.. European Journal of Biochemistry 209:367–374
    [Google Scholar]
  10. Michaelis L., Gramick S. 1943; The polymerization of the free radicals of the Wurster dye type: the dimeric resonance band.. Journal of the American Chemical Society 65:1747–1755
    [Google Scholar]
  11. Okuzumi M., Kita Y. 1965; Studies on biochemistry of the thiobacilli. VI. Oxidation of thiosulphate to tetrathionate by T. thiooxidans.. Agricultural and Biological Chemistry 29:1063–1068
    [Google Scholar]
  12. Pronk J.T., Meuleneerg R., Hazeu W., Bos P., Kuenen J.G. 1990a; Oxidation of reduced inorganic sulphur compounds by acidophilic thiobacilli.. FEMS Microbiology Reviews 75:293–306
    [Google Scholar]
  13. Pronk J.T., Meesters P.J.W., Van DIJKEN J.P., Bos P., Kuenen J.G. 1990b; Heterotrophic growth of Thiobacillus acidophilus in batch and chemostat cultures. Archices of Microhiology 153:392–398
    [Google Scholar]
  14. Pronk J.T., Meulenberg R., Van DEN BERG D.J.C., Batenburg VAN DER VEGTE W., Bos P., Kuenen J.G. 1990c; Heterotrophic growth of Thiobacillus acidophilus in batch and chemostat cultures. Applied and Environmental Microbiology 56:3395–3401
    [Google Scholar]
  15. Rieske J.S. 1967; he quantitative determination of mitochondria1 hemoproteins.. Methods in Enzymology 10:488–493
    [Google Scholar]
  16. Roy A.B., Trudinger P.A. 1970 The Biochemistry of Inorganic Compounds of Sulphur. Cambridge: University Press;
    [Google Scholar]
  17. Silver M., Lundgren D. G. 1968; .The thiosulfate-oxidizing enzyme of Ferrobacillus ferrooxidans (Thiobacillus ferrooxidans).. Canadian Journal of Biochemistry 46:1215–1220
    [Google Scholar]
  18. Sinha D.B., Walden C.C. 1966; .ormation of polythionates and their interrelationships during oxidation of thiosulphate by T. -ferrooxidans.. Canadian Journal of Biochemistry 12:1041–1054
    [Google Scholar]
  19. S�RBO B. 1957; A colorimetric method for the determination of thiosulphate.. Biochimica et Biophysica Acta 23:412–416
    [Google Scholar]
  20. Thomas P.E., Ryan D., Levin W. 1976; An improved staining procedure for the detection of the peroxidase activity of cytochrome P450 on sodium dodecyl sulfate polyacrylamide gels.. Analytical Biochemistry 75:168–176
    [Google Scholar]
  21. Trudinger P.A. 1961; Thiosulphate oxidation and cytochromes in Thiobacillus X.. Biochemical Journal 78:680–686
    [Google Scholar]
  22. Vaniersel J., Frank JZN J., Duine J.A. 1985; Determination of absorption coefficients of purified proteins by conventional ultraviolet spectrophotometry and chromatography combined with multiwavelength detection.. Analytical Biochemistry 151:196–204
    [Google Scholar]
  23. Wood A.P., Kelly D.P. 1986; Chemolithotrophic metabolism of the newly-isolated moderately thermophilic, obligately autotrophic Thiobacillus tepidarius.. Archives of Microbiology 144:71–77
    [Google Scholar]
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