1887

Microbiology Society logo

Volume 131, Issue 2

Glucose-6-phosphate Dehydrogenase from : Purification and Properties Free

Abstract

glucose-6-phosphate dehydrogenase (EC 1.1.1.49) was purified to homogeneity. The subunit molecular weight was 55300 and the molecular weight of the native protein was 138000 by gel filtration and 131 800 by rate zonal ultracentrifugation on sucrose gradients. There was some evidence for a small population of higher molecular weight oligomers during rate zonal centrifugation. A sedimentation coefficient of 8·5S was determined by analytical ultracentrifugation (suggesting the presence of the tetramer). Three bands of enzyme activity were separated on polyacrylamide gels. Digests of the proteins in the three bands gave peptide patterns with no detectable difference after SDS electrophoresis. Linear double reciprocal plots were obtained with glucose 6-phosphate as a variable substrate. The apparent K for glucose 6-phosphate was 214 μ and the was 520 μmol (mg protein) min. A sigmoidal saturation curve and a non-linear double reciprocal plot were observed for NADP, giving a Hill constant of 1·5 and [S] for NADP of 89 μ. The enzyme was inhibited by NADPH, with 50% inhibition at a NADP: NADPH ratio of 0·4 when total NADP(H) concentration was 0·2 m. ATP, ADP, AMP and cAMP had no effect on enzyme activity. Palmitoyl CoA inhibited the initial velocity of the reaction by 95% at a concentration of 1·5 μ when incubated with enzyme prior to NADP addition.

  • Received:
  • Revised:
  • Published Online:
© Society for General Microbiology 1996
Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-131-2-321
1985-02-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/micro/131/2/mic-131-2-321.html?itemId=/content/journal/micro/10.1099/00221287-131-2-321&mimeType=html&fmt=ahah

References

  1. Ashihara H., Komamine A. 1974; Regulation of the activities of some enzymes of the pentose phosphate pathway in Phaseolus mungo. Zeitschrift für Pflanzenphysiologie 74:130–142
     [Google Scholar]
  2. Axelsen N. H., Kroll J., Weeke B. 1973; A Manual of Quantitive Electrophoresis. Oslo: Universitetsforlaget;
     [Google Scholar]
  3. Bonsignore A., Cancedda R., Nicolini A., Da-miani G., DeFlora A. 1971; Metabolism of human erythrocyte glucose-6-phosphate dehydro-genase. VI Interconversion of multiple molecular forms. Archives of Biochemistry and Biophysics 147:493–501
     [Google Scholar]
  4. Bradford M. 1976; A rapid and sensitive method for the quantitation of microgram quantities of protein using the principle of protein-dye binding. Analytical Biochemistry 71:248–254
     [Google Scholar]
  5. Cebrian-Perez J. A., Garcia-Gimeno M. A., Muino-Blanco T. 1980; Isoenzimas de la glucosa-6-fosfato deshidrogenasa de Aspergillus oryzae (Ahlburg). Revista espahola defisiologia 36:389–394
     [Google Scholar]
  6. Davis B. 1964; Disc electrophoresis. II Method and applications to human serum proteins. Annals of the New York Academy of Sciences 121:404–427
     [Google Scholar]
  7. Donohue T. M., Mahowald T. A., Adams D. J., Barker K. L. 1981; Glucose-6-phosphate dehy-drogenase. Partial characterization of the rat liver and uterine enzymes. Biochimica et biophysica acta 658:356–368
     [Google Scholar]
  8. Dütsch G. A., Rast D. 1972; Biochemische Beziehung zwischen Mannitbildung und Hexose-monophosphatzyklus in Agaricus bisporus. Phytoche-mistry 11:2677–2681
     [Google Scholar]
  9. Edmundowicz J. M., Wriston J. C Jr. 1963; Mannitol dehydrogenase from Agaricus campestris. Journal of Biological Chemistry 238:3539–3541
     [Google Scholar]
  10. Eggleston L. V., Krebs H. A. 1974; Regulation of the pentose phosphate cycle. Biochemical Journal 138:425–435
     [Google Scholar]
  11. Engel H. J., Domschke W., Alberti M., Domagk G. F. 1969; Protein structure and enzymatic activity. II Purification and properties of a crystalline glucose-6-phosphate dehydrogenase from Candida utilis. Biochimica et biophysica acta 191:509–516
     [Google Scholar]
  12. Hammond J. B. W. 1977; Carbohydrate metabolism in Agaricus bisporus: oxidative pathways in mycelium and sporophore. Journal of General Microbiology 102:245–248
     [Google Scholar]
  13. Hammond J. B. W. 1981; Variations in enzyme activity during periodic fruiting of Agaricus bisporus. New Phytologist 89:419–428
     [Google Scholar]
  14. Hammond J. B. W. 1985; Sugar, sugar phosphate and NADP(H) levels in Agaricus bisporus fruit bodies. Journal of General Microbiology 131:329–333
     [Google Scholar]
  15. Hammond J. W., Nichols R. 1976; Carbohydrate metabolism in Agaricus bisporus (Lange) Sing.: changes in soluble carbohydrates during growth of mycelium and sporophore. Journal of General Microbiology 93:309–320
     [Google Scholar]
  16. Hou H. H., Wu L. C., Chen C. C. 1975; Respiratory pathways of the cultivated mushroom, Agaricus bisporus. Mushroom Science 9:37–49
     [Google Scholar]
  17. Kawaguchi A., Bloch K. 1974; Inhibition of glucose 6-phosphate dehydrogenase by palmitoyl CoA. Journal of Biological Chemistry 249:5793–5800
     [Google Scholar]
  18. Kurganov B. 1982; Allosteric Enzymes - Kinetic Behaviour. Chichester, UK: John Wiley & Sons;
     [Google Scholar]
  19. Le roux P. 1967; Importance relative de la voie des pentoses phosphates et de la glycolyse dans le carpophore d’Agaricus bisporus. Annales de Physiologic végetale 9:349–363
     [Google Scholar]
  20. Levy H. R. 1979; Glucose-6-phosphate dehydrogen-ases. Advances in Enzymology 48:97–192
     [Google Scholar]
  21. Levy H. R., Christoff M. 1983; A critical appraisal of the effect of oxidized glutathione on hepatic glucose-6-phosphate dehydrogenase activity. Biochemical Journal 214:959–965
     [Google Scholar]
  22. Lischwe M. A., Ochs D. 1982; A new method for partial peptide mapping using . N-chlorosuccini-mide/urea and peptide silver staining in sodium dodecyl sulfate-polyacrylamide gels. Analytical Biochemistry 127:453–457
     [Google Scholar]
  23. Luzzato L. 1967; Regulation of the activity of glucose-6-phosphate dehydrogenase by NADP and NADPH. Biochimica et biophysica acta 146:18–25
     [Google Scholar]
  24. Markham R. 1960; A graphical method for the rapid determination of sedimentation coefficients. Biochemical Journal 77:516–519
     [Google Scholar]
  25. Martin R. A., Ames B. N. 1961; A method for determining the sedimentation behaviour of enzymes. Application to protein mixtures. Journal of Biological Chemistry 236:1372–1379
     [Google Scholar]
  26. Mita M., Yasumasu I. 1980; Inhibition of glucose-6-phosphate dehydrogenase and 6-phosphoglucon-ate dehydrogenase in sea urchin eggs by palmitoyl coenzyme A and reversal by polyamines. Archives of Biochemistry and Biophysics 201:322–329
     [Google Scholar]
  27. Muto S., Uritani I. 1971; Effect of NADP and glucose-6-phosphate on the sedimentation behaviour of glucose-6-phosphate dehydrogenase from sweet potato. Plant and Cell Physiology 12:803–806
     [Google Scholar]
  28. Noltmann E. A., Gubler C. J., Kuby S. A. 1961; Glucose 6-phosphate dehydrogenase (Zwischenfer-ment). 1. Isolation of the crystalline enzyme from yeast. Journal of Biological Chemistry 236:1225–1230
     [Google Scholar]
  29. Opheim D., Bernlohr R. W. 1973; Purification and regulation of glucose-6-phosphate dehydrogenase from Bacillus licheniformis. Journal of Bacteriology 116:1150–1159
     [Google Scholar]
  30. Ouchterlony O. 1949; Antigen-antibody reactions in gels. Acta pathologica et microbiologica scandina-vica 26:507–515
     [Google Scholar]
  31. Taketa K., Pogell B. M. 1966; The effect of palmitoyl coenzyme A on glucose-6-phosphate dehydrogenase and other enzymes. Journal of Biological Chemistry 241:720–726
     [Google Scholar]
  32. Veech R. L., Eggleston L. V., Krebs H. A. 1969; The redox state of free nicotinamide adenine dinucleotide phosphate in the cytoplasm of rat liver. Biochemical Journal 115:609–619
     [Google Scholar]
  33. Weber K., Osborne M. 1969; Reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide-gel electrophoresis. Journal of Biological Chemistry 244:4406–4412
     [Google Scholar]
  34. Wood D. 1980; Production, purification and properties of extracellular laccase of Agaricus bisporus. Journal of General Microbiology 111:327–338
     [Google Scholar]
  35. Wray W., Boulikas T., Wray V. P., Hancock R. 1981; Silver staining of proteins in polyacrylamide gels. Analytical Biochemistry 118:197–203
     [Google Scholar]
  36. Yielding K. L. 1970; Modification by sucrose of the catalytic activity and physical properties of glutamic dehydrogenase. Biophysical and Biochemical Research Communications 38:546–551
     [Google Scholar]
  37. Yue R. H., Noltmann E. A., Kuby S. A. 1969; Glucose-6-phosphate dehydrogenase from brewer’s yeast (zwischenferment). III Studies on the sub-unit structure and on the molecular association phenomenon induced by triphosphoryridine nucleo-tide. Journal of Biological Chemistry 244:1353–1364
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-131-2-321
Loading
Glucose-6-phosphate Dehydrogenase from Agaricus bisporus: Purification and Properties
Microbiology 131, 321 (1985); https://doi.org/10.1099/00221287-131-2-321
/content/journal/micro/10.1099/00221287-131-2-321
/content/journal/micro/10.1099/00221287-131-2-321
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error