1887

Abstract

Using a continuous culture of MGA3 limited by 100 mM methanol in the feed and growing at a dilution rate =025 h, transients in dissolved methanol were studied to determine the effects of methanol toxicity and the pathway of methanol dissimilation to CO. Steady-state cultures were disturbed by pulses of methanol resulting in a rapid change in concentration of 64–128 mM. MGA3 responded to a sudden increase in available methanol by a transient decline in the biomass concentration in the reactor. In most cases the culture returned to steady state between 4 and 12 h after pulse addition. However, at a methanol pulse of 128 mM, complete biomass washout occurred and the culture did not return to steady state. Integrating the response curves of the dry biomass concentration over a 12 h time period showed that a methanol pulse can cause an average transient decline in the biomass yield of up to 22%. C NMR experiments using labelled methanol indicated that the transient partial or complete biomass washout was probably caused by toxic accumulation of formaldehyde in the culture. These experiments also showed accumulation of formate, indicating that possesses formaldehyde dehydrogenase and formate dehydrogenase activity resulting in a methanol dissimilation pathway via formate to CO. Studies using isotope-ratio mass spectrometry provided further evidence of a methanol dissimilation pathway via formate. MGA3, growing continuously under methanol limitation, consumed added formate at a rate of approximately 085 mmol l h. Furthermore, significant accumulation of CO in the reactor exhaust gas was measured in response to a pulse addition of [C]formic acid to the bioreactor. This indicates that dissimilates methanol carbon to CO in order to detoxify formaldehyde by both a linear pathway to formate and a cyclic mechanism as part of the RuMP pathway.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-148-10-3223
2002-10-01
2020-09-21
Loading full text...

Full text loading...

/deliver/fulltext/micro/148/10/1483223a.html?itemId=/content/journal/micro/10.1099/00221287-148-10-3223&mimeType=html&fmt=ahah

References

  1. Al-Awadhi N, Egli T., Hamer G. 1988; Growth characteristics of a thermotolerant methylotrophic Bacillus sp. (NCIB; 12522) in batch culture. Appl Microbiol Biotechnol29:485–493
    [Google Scholar]
  2. Al-Awadhi N, Egli T, Hamer G., Mason C. A. 1990; The process utility of thermotolerant methylotrophic bacteria: II. An evaluation of transient responses. Biotechnol Bioeng36:821–825
    [Google Scholar]
  3. Anthony C. 1978; The prediction of growth yields in methylotrophs. J Gen Microbiol104:91–104
    [Google Scholar]
  4. Anthony C. 1982; The Biochemistry of Methylotrophs London: Academic Press;
    [Google Scholar]
  5. Arfman N, Watling E. M, Clement W, van Oosterwijk R. J, de Vries G. E, Harder W, Attwood M. M., Dijkhuizen L. 1989; Methanol metabolism in thermotolerant methylotrophic Bacillus strains involving a novel catabolic NAD-dependent methanol dehydrogenase as a key enzyme. Arch Microbiol152:280–288
    [Google Scholar]
  6. Arfman N, van Beeumen J, de Vries G. E, Harder W., Dijkhuizen L. 1991; Purification and characterization of an activator protein for methanol dehydrogenase from thermotolerant Bacillus spp. J Biol Chem266:3955–3960
    [Google Scholar]
  7. Arfman N, Dijkhuizen L, Kirchhof G.. 8 other authors 1992; Bacillus methanolicus sp. nov., a new species of thermotolerant, methanol-utilizing, endospore-forming bacteria. Int J Syst Bacteriol42:439–445
    [Google Scholar]
  8. Bacher A, Rieder C, Eichinger D, Arigoni D, Fuchs G., Eisenreich W. 1998; Elucidation of novel biosynthetic pathways and metabolite flux patterns by retrobiosynthetic NMR analysis. FEMS Microbiol Rev22:567–598
    [Google Scholar]
  9. Beyer H., Walter W. 1988; Lehrbuch der Organischen Chemie, 21st edn. Stuttgart: S. Hirzel Verlag;
    [Google Scholar]
  10. Brecker L., Ribbons D. W. 2000; Biotransformations monitored in situ by proton nuclear magnetic resonance spectroscopy. Trends Biotechnol18:197–202
    [Google Scholar]
  11. Brecker L, Urdl P, Schmid W, Griengl H., Ribbons D. W. 2000; Simple device to monitor aerobic biotransformations by in situ 1H-NMR. Biotechnol Lett22:1135–1141
    [Google Scholar]
  12. Brooke A. G, Watling E. M, Attwood M. M., Tempest D. W. 1989; Environmental control of metabolic fluxes in thermotolerant methylotrophic Bacillus strains. Arch Microbiol151:268–273
    [Google Scholar]
  13. Brooke A. G, Attwood M. M., Tempest D. W. 1990; Metabolic fluxes during the growth of thermotolerant methylotrophic Bacillus strains in methanol-sufficient chemostat cultures. Arch Microbiol153:591–595
    [Google Scholar]
  14. Chistoserdova L, Gomelsky L, Vorholt J. A, Gomelsky M, Tsygankov Y. D., Lidstrom M. E. 2000; Analysis of two formaldehyde oxidation pathways in Methylobacillus flagellatus KT, a ribulose monophosphate cycle methylotroph. Microbiology146:233–238
    [Google Scholar]
  15. Christensen B., Nielsen J. 2002; Reciprocal 13C-labeling: a method for investigating the catabolism of cosubstrates. Biotechnol Progr18:163–166
    [Google Scholar]
  16. Cornish A, Nicholls K. M, Scott D, Hunter B. K, Aston W. J, Higgins I. J., Sanders J. K. 1984; In vivo 13C NMR investigations of methanol oxidation by the obligate methanotroph Methylosinus trichosporium OB3b. J Gen Microbiol130:2565–2575
    [Google Scholar]
  17. Dijkhuizen L, Arfman N, Attwood M. M, Brooke A. G, Harder W., Watling E. M. 1988; Isolation and initial characterization of methylotrophic Bacillus strains. FEMS Microbiol Lett52:209–214
    [Google Scholar]
  18. De Vries G. S, Arfman N, Terpstra P., Dijkuizen L. 1992; Cloning, expression, and sequence analysis of the Bacillus methanolicus C1 methanol dehydrogenase gene. J Bacteriol174:5346–5353
    [Google Scholar]
  19. Evans C. G. T, Herbert D., Tempest D. W. 1970; The continuous cultivation of micro-organisms: 2. Construction of a chemostat. Methods Microbiol2:277–327
    [Google Scholar]
  20. Gı́rio F. M, Amaral-Collago M. T., Attwood M. M. 1995; Physiological responses of a methylotrophic bacterium after sudden shifts from C-limited chemostat to C-excess batch growth conditions. J Appl Bacteriol79:409–416
    [Google Scholar]
  21. Hanson R. S, Dillingham R, Olson P, Lee G. H, Cue D, Schendel F. J, Bremmon C., Flickinger M. C. 1996; Production of l-lysine and some other amino acids by mutants of Bacillus methanolicus . In Microbial Growth on C1 Compounds: Proceedings of the 7th International Symposium on Microbial Growth on C1 Compounds pp227–236 Edited by Lidstrom M. E., Tabita F. R.. Dordrecht: Kluwer;
    [Google Scholar]
  22. Hunter B. K, Nicholls K. M., Sanders J. K. 1984; Formaldehyde metabolism by E. coli . In vivo carbon, deuterium, and two-dimensional NMR observations of multiple detoxifying pathways. Biochemistry23:508–514
    [Google Scholar]
  23. Johnson B. J, Borowski J., Engbolm C. 1964; Steam sterilizable probes for dissolved oxygen measurement. Biotechnol Bioeng6:457–468
    [Google Scholar]
  24. Jones J. G., Bellion E. 1991; Methanol oxidation and assimilation in Hansenula polymorpha . An analysis by 13C NMR in vivo . Biochem J280:475–481
    [Google Scholar]
  25. Leak D. J. 1992; Biotechnological and applied aspects of methane and methanol utilizers. In Methane and Methanol Utilizers pp245–279 Edited by Murrell J. C., Dalton H.. New York: Plenum Press;
    [Google Scholar]
  26. Leak D. J. 1999; Applications of methylotrophs. In The Encyclopedia of Bioprocess Technology: Fermentation, Biocatalysis, and Bioseparation pp1742–1753 Edited by Flickinger M. C., Drew S. W.. New York: Wiley;
    [Google Scholar]
  27. Lee G. H, Hur W, Bremmon C. E., Flickinger M. C. 1996; Lysine production from methanol at 50 °C using Bacillus methanolicus : modeling volume control, lysine concentration, and productivity using a three-phase continuous simulation. Biotechnol Bioeng49:639–653
    [Google Scholar]
  28. Levy G. C, Lichter R. L., Nelson G. L. 1980; Carbon-13 Nuclear Magnetic Resonance Spectroscopy New York: Wiley;
    [Google Scholar]
  29. Marchenko G. N, Marchenko N. D, Tsygankov Y. D., Chistoserdova A. Y. 1999; Organization of threonine biosynthetic genes from the obligate methylotroph Methylobacillus flagellatus . Microbiology145:3273–3282
    [Google Scholar]
  30. Pluschkell S. B. 1999; Characterization and mathematical modeling of growth and glutamic acid production by Bacillus methanolicus MGA3. Diss Abstr Int B Sci Eng59:6404
    [Google Scholar]
  31. Rehmann K, Hertkorn N., Kettrup A. A. 2001; Fluoranthene metabolism in Mycobacterium sp. strain KR20: identity of pathway intermediates during degradation and growth. Microbiology147:2783–2794
    [Google Scholar]
  32. Schendel F. J, Bremmon C. E, Flickinger M. C, Guettler M., Hanson R. S. 1990; l-Lysine production at 50 °C by mutants of a newly isolated and characterized methylotrophic Bacillus sp. Appl Environ Microbiol56:963–970
    [Google Scholar]
  33. Shimizu K. 2000; An overview on metabolic systems engineering approach and its future perspectives for efficient microbial fermentation. J Chin Inst Chem Eng30:429–442
    [Google Scholar]
  34. Vonk J, Arfman N, de Vries G. E, van Beeumen J, van Bruggen E. F., Dijkhuizen L. 1991; Electron microscopic analysis and biochemical characterization of a novel methanol dehydrogenase from the thermotolerant Bacillus sp. C1. J Biol Chem 266:3949–3954
    [Google Scholar]
  35. Vorholt J. A, Chistoserdova L, Stolyar S. M, Thauer R. K., Lidstrom M. E. 1999; Distribution of tetrahydromethanopterin-dependent enzymes in methylotrophic bacteria and phylogeny of methenyl tetrahydromethanopterin cyclohydrolases. J Bacteriol181:5750–5757
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-148-10-3223
Loading
/content/journal/micro/10.1099/00221287-148-10-3223
Loading

Data & Media loading...

Most cited this month 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