Skip to content
1887

Microbiology Society logo Microbiology

Volume 107, Issue 1

Purification and Properties of an NAD(P)-linked Formaldehyde Dehydrogenase from (Bath) Free

Abstract

Crude soluble extracts of strain Bath, grown on methane, were found to contain NAD(P)-linked formaldehyde dehydrogenase activity. Activity in the extract was lost on dialysis against phosphate buffer, but could be restored by supplementing with inactive, heat-treated extract (70 °C for 12 min). The non-dialysable, heat-sensitive component was isolated and purified, and has a molecular weight of about 115000. Sodium dodecyl sulphate gel electrophoresis of the protein suggested there were two equal subunits with molecular weights of 57000. The heat-stable fraction, which was necessary for activity of the heat-sensitive protein, was trypsin-sensitive and presumed to be a low molecular weight protein or peptide. A number of thiol compounds and other common cofactors could not replace the component present in the heat-treated soluble extract. The purified formaldehyde dehydrogenase oxidized three other aldehydes with the following values: 0·68 m (formaldehyde); 0·075 m (glyoxal); 7·0 m (glycolaldehyde); and 2·0 m (-glyceraldehyde). NAD or NADP was required for activity, with values of 0.063 and 0.155 m respectively, and could not be replaced by any of the artificial electron acceptors tested. The enzyme was heat-stable at 45 °C for at least 10 min and had temperature and pH optima of 45 °C and pH 7.2 respectively. A number of metal-binding agents and substrate analogues were not inhibitory. Thiol reagents gave varying degrees of inhibition, the most potent being -hydroxymercuribenzoate which at 1 m gave 100 % inhibition. The importance of possessing an NAD(P)-linked formaldehyde dehydrogenase, with respect to , is discussed.

  • Received:
  • Published Online:
© Society for General Microbiology 1978
Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-107-1-19
1978-07-01
2025-04-20
Loading full text...

Full text loading...

/deliver/fulltext/micro/107/1/mic-107-1-19.html?itemId=/content/journal/micro/10.1099/00221287-107-1-19&mimeType=html&fmt=ahah

References

  1. Andrews P. 1964; Estimation of the molecular weight of proteins by Sephadex gel-filtration. Biochemical Journal 91:222–233
     [Google Scholar]
  2. Anthony C. 1978; The prediction of growth yields in methylotrophs. Journal of General Microbiology 104:91–104
     [Google Scholar]
  3. Anthony C., Zatman L. J. 1964; The microbial oxidation of methanol. 2. The methanol-oxidizing enzyme of Pseudomonas sp. m27. Biochemical Journal 92:614–621
     [Google Scholar]
  4. Boulton C. A., Large P. J. 1977; Synthesis of certain assimilatory and dissimilatory enzymes during bacterial adaptation to growth on trimethyl-amine. Journal of General Microbiology 101:151–156
     [Google Scholar]
  5. Chrastil J., Wilson J. A. 1975; A sensitive colorimetric method for formaldehyde. Analytical Biochemistry 63:202–207
     [Google Scholar]
  6. Colby J., Dalton H. 1976; Some properties of a soluble methane mono-oxygenase from Methylococcus capsulatus strain Bath. Biochemical Journal 157:495–197
     [Google Scholar]
  7. Colby J., Zatman L. J. 1973; Trimethylamine metabolism in obligate and facultative methylotrophs. Biochemical Journal 132:101–112
     [Google Scholar]
  8. Colby J., Zatman L. J. 1975; Enzymological aspects of the pathways for trimethylamine oxidation and C1 assimilation in obligate methylotrophs and restricted facultative methylotrophs. Biochemical Journal 148:513–520
     [Google Scholar]
  9. Colby J., Stirling D. I., Dalton H. 1977; The soluble methane mono-oxygenase of Methylococcus capsulatus (Bath). Its ability to oxygenate n-alkanes, n-alkenes, ethers and alicyclic, aromatic and heterocyclic compounds. Biochemical Journal 165:395–402
     [Google Scholar]
  10. Dalton H. 1977; Ammonia oxidation by the methane oxidising bacterium Methylococcus capsulatus strain Bath. Archives of Microbiology 114:273–279
     [Google Scholar]
  11. Dalton H., Whittenbury R. 1976; The acetylene reduction technique as an assay for the nitro-genase activity in the methane oxidizing bacterium Methylococcus capsulatus strain Bath. Archives of Microbiology 109:147–151
     [Google Scholar]
  12. Eisenthal R., Cornish-Bowden A. 1974; The direct linear plot. A new graphical procedure for estimating enzyme kinetic parameters. Biochemical Journal 139:715–720
     [Google Scholar]
  13. Frisell W. R., Mackenzie C. G. 1963; The determination of formaldehyde and serine in biological systems. Methods of Biochemical Analysis 6:, 2.63–77
     [Google Scholar]
  14. Gabriel O. 1971; Analytical disc gel electro-phoresis. Methods in Enzymology 22:565–578
     [Google Scholar]
  15. Hampton D., Zatman L. J. 1973; The metabolism of tetramethylammonium chloride by Bacterium 5h2. Biochemical Society Transactions 1:667–668
     [Google Scholar]
  16. Harrington A. A., Kallio R. E. 1960; Oxidation of methanol and formaldehyde by Pseudomonas methanica . Canadian Journal of Microbiology 6:1–7
     [Google Scholar]
  17. Johnson P. A., Quayle J. R. 1964; Microbial growth on C1 compounds. 6. Oxidation of methanol, formaldehyde and formate by methanol-grown Pseudomonas amI. Biochemical Journal 93:281–290
     [Google Scholar]
  18. Kung H. F., Wagner C. 1970; Oxidation of C1 compounds by Pseudomonas sp. ms. Biochemical Journal 116:357–365
     [Google Scholar]
  19. Lang E., Lang H. 1972; Spezifische Farbreaktion zum direkten Nachweis der Ameisensäure. Zeitschrift für analytische Chemie 260:8–10
     [Google Scholar]
  20. Large P. J., Quayle J. R. 1963; Microbial growth on C1 compounds. 5. Enzyme activities in extracts of Pseudomonas amI. Biochemical Journal 87:386–396
     [Google Scholar]
  21. Lowry O. H., Rosebrough N. J., Farr A. L., Randall R. J. 1951; Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry 193:265–275
     [Google Scholar]
  22. Mehta R. J. 1973; Studies on methanol-oxidizing bacteria. II. Purification and properties of methanol dehydrogenase from Pseudomonas rj 1 . Antonie van Leeuwenhoek 39:303–312
     [Google Scholar]
  23. Mehta R. J. 1975; A novel inducible formaldehyde dehydrogenase of Pseudomonas sp. (rj 1). Antonie van Leeuwenhoek 41:89–95
     [Google Scholar]
  24. Nash T. 1953; The colorimetric estimation of formaldehyde by means of the Hantzsch reaction. Biochemical Journal 55:416–421
     [Google Scholar]
  25. O’Connor M. L., Hanson R. S. 1977; Enzyme regulation in Methylobacterium organophilum . Journal of General Microbiology 101:327–332
     [Google Scholar]
  26. Patel R. N., Felix A. 1976; Microbial oxida tion of methane and methanol : crystallization and properties of methanol dehydrogenase from Methylosinus sporium . Journal of Bacteriology 128:413–424
     [Google Scholar]
  27. Patel R. N., Hoare D. S. 1971; Physiological studies of methane and methanol-oxidizing bacteria: oxidation of C-1 compounds by Methylococcus capsuiatus . Journal of Bacteriology 107:187–192
     [Google Scholar]
  28. Sperl G. T., Forrest H. S., Gibson D. T. 1974; Substrate specificity of the purified primary alcohol dehydrogenase from methanol-oxidizing bacteria. Journal of Bacteriology 178:541–550
     [Google Scholar]
  29. Stirling D. I., Dalton H. 1977; Effect of metal-binding agents and other compounds on methane oxidation by two strains of Methylococcus capsuiatus . Archives of Microbiology 114:71–76
     [Google Scholar]
  30. Strøm T., Ferenci T., Quayle J. R. 1974; The carbon assimilation pathways of Methylococcus capsuiatus Pseudomonas methanica and Methylosinus trichosporium (ob3b) during growth on methane. Biochemical Journal 144:465–476
     [Google Scholar]
  31. Tonge G. M., Harrison D. E. F., Higgins I. J. 1977; Purification and properties of the methane mono-oxygenase enzyme system from Methylosinus trichosporium oe3b . Biochemical Journal 161:333–344
     [Google Scholar]
  32. Trudinger P. A. 1969; Thiol reagents. Data for Biochemical Research, 2.388–395 Dawson R. M. C., Elliot D. C., Elliot W. H., Jones K. M. London: Oxford University Press;
     [Google Scholar]
  33. Van Dijken J. P., Harder W. 1975; Growth yields of micro-organisms on methanol and methane. A theoretical study. Biotechnology and Bioengineering 17:15–30
     [Google Scholar]
  34. Van Dijken J. P., Oostra-Demkes G. T., Otto R., Harder W. 1976; S-Formylglutathione : the substrate for formate dehydrogenase in methanol-utilizing yeasts. Archives of Microbiology 111:77–83
     [Google Scholar]
  35. Whittenbury R., Phillips K. C., Wilkinson J. F. 1970; Enrichment, isolation and some properties of methane-utilizing bacteria. Journal of General Microbiology 61:205–218
     [Google Scholar]
/content/journal/micro/10.1099/00221287-107-1-19
Loading
Purification and Properties of an NAD(P)+-linked Formaldehyde Dehydrogenase from Methylococcus capsulatus (Bath)
Microbiology 107, 19 (1978); https://doi.org/10.1099/00221287-107-1-19
/content/journal/micro/10.1099/00221287-107-1-19
/content/journal/micro/10.1099/00221287-107-1-19
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