Applied Microbiology and Biotechnology

, Volume 93, Issue 4, pp 1777–1784 | Cite as

Metabolic control of Clostridium thermocellum via inhibition of hydrogenase activity and the glucose transport rate

  • Hsin-Fen Li
  • Barbara L. Knutson
  • Sue E. Nokes
  • Bert C. Lynn
  • Michael D. Flythe
Bioenergy and biofuels


Clostridium thermocellum has the ability to catabolize cellulosic biomass into ethanol, but acetic acid, lactic acid, carbon dioxide, and hydrogen gas (H2) are also produced. The effect of hydrogenase inhibitors (H2, carbon monoxide (CO), and methyl viologen) on product selectivity was investigated. The anticipated effect of these hydrogenase inhibitors was to decrease acetate production. However, shifts to ethanol and lactate production are also observed as a function of cultivation conditions. When the sparge gas of cellobiose-limited chemostat cultures was switched from N2 to H2, acetate declined, and ethanol production increased 350%. In resting cell suspensions, lactate increased when H2 or CO was the inhibitor or when the cells were held at elevated hyperbaric pressure (6.8 atm). In contrast, methyl-viologen-treated resting cells produced twice as much ethanol as the other treatments. The relationship of chemostat physiology to methyl viologen inhibition was revealed by glucose transport experiments, in which methyl viologen decreased the rate of glucose transport by 90%. C. thermocellum produces NAD+ from NADH by H2, lactate, and ethanol production. When the hydrogenases were inhibited, the latter two products increased. However, excess substrate availability causes fructose 1,6-diphosphate, the glycolytic intermediate that triggers lactate production, to increase. Compensatory ethanol production was observed when the chemostat fluid dilution rate or methyl viologen decreased substrate transport. This research highlights the complex effects of high concentrations of dissolved gases in fermentation, which are increasingly envisioned in microbial applications of H2 production for the conversion of synthetic gases to chemicals.


Chemostat Consolidated bioprocessing Cellulosic biomass conversion Cellulose Metabolic engineering Product selectivity Continuous culture Paraquat 



The authors gratefully acknowledge the financial support of the Southeastern Sun Grant Center, administered by the University of Tennessee, grant number DTOS59-07-G-0050. MF was supported by the Agricultural Research Service, USDA. The authors would like to thank Gloria Gellin and Jerry Vice for the technical assistance.


“Proprietary or brand names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product, nor exclusion of others that may be suitable.”


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Copyright information

© Springer-Verlag (outside the USA) 2011

Authors and Affiliations

  • Hsin-Fen Li
    • 1
  • Barbara L. Knutson
    • 1
  • Sue E. Nokes
    • 2
  • Bert C. Lynn
    • 3
  • Michael D. Flythe
    • 4
    • 5
  1. 1.Department of Chemical and Materials EngineeringUniversity of KentuckyLexingtonUSA
  2. 2.Department of Biosystems and Agricultural EngineeringUniversity of KentuckyLexingtonUSA
  3. 3.Department of ChemistryUniversity of KentuckyLexingtonUSA
  4. 4.Forage-Animal Production Research Unit, Agricultural Research ServiceUSDALexingtonUSA
  5. 5.Department of Animal and Food SciencesUniversity of KentuckyLexingtonUSA

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