Abstract
Ethanol inhibition of glucose catabolism in Zymomonas mobilis was investigated using 31P NMR spectroscopy in vivo and of perchloric acid extracts from cell suspensions incubated with 0, 5 and 10% (w/v) ethanol. In vivo 31P NMR experiments revealed slower glucose utilization and decreased levels of nucleoside triphosphates in the presence of 10% ethanol as compared to controls. Using 31P NMR spectroscopy of perchloric acid extracts, intracellular accumulation of 3.4 mM 3-phosphoglycerate was found when 10% ethanol was present in the medium. No accumulation of this metabolite occurred in cells incubated with 0 and 5% ethanol. Enzyme assays confirmed that phosphoglycerate-mutase and enolase were inhibited 31 and 40%, respectively, in the presence of 10% ethanol in the test system. Therefore, under the conditions used the decrease in the fermentative activity of Z. mobilis at high ethanol concentrations is due to inhibition of phosphoglycerate-mutase and enolase.
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Abbreviations
- KDPG:
-
2-keto-3-deoxy-6-phosphogluconate
References
AlgarEM, ScopesRK (1985) Studies on cell-free metabolism: ethanol production by extracts of Zymomonas mobilis. J Biotechnol 2: 275–287
BarrowKD, CollinsJG, NortonRS, RogersPL, SmithGM (1984) 31P Nuclear magnetic resonance studies of the fermentation of glucose to ethanol by Zymomonas mobilis. J Biol Chem 259: 5711–5716
BergmeyerHU, BergmeyerJ, GrasslM (1984) Methods of enzymatic analysis, vol 2. Verlag Chemie, Weinheim, pp 182–183
BradfordMM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254
BringerS, FinnRK, SahmH (1984) Effect of oxygen on the metabolism of Zumomonas mobilis. Arch Microbiol 139: 376–381
CareyVC, IngramLO (1983) Lipid composition of Zymomonas mobilis: effects of ethanol and glucose. J Bacteriol 154: 1291–1300
DenHollanderJA, UgurbilK, BrownTR, ShulmanRG (1981) Phosphorus-31 nuclear magnetic resonance studies of the effect of oxygen upon glycolysis in yeast. Biochemistry 20: 5871–5880
FeldmannSD, SahmH, SprengerGA (1992) Pentose metabolism in Zymomonas mobilis wild type and recombinant strains. Appl Microbiol Biotechnol 38: 354–361
HermansMAF, NeussB, SahmH (1991) Content and composition of hopanoids in Zymomonas mobilis under various growth conditions. J Bacteriol 173: 5592–5595
IngramLO, ButtkeTM (1984) Effects of alcohols on microorganisms Adv Microb Physiol 25: 253–300
KirkK, RaftosJE, KuchelPW (1986) Triethyl phosphate as an internal 31P NMR reference in biological samples. J Magn Reson 70: 484–487
LeeKJ, SkotnickiML, TribeDE, RogersPL (1980) Kinetic studies on a highly productive strain of Zymomonas mobilis. Biotechnol Lett 2: 339–344
Loureiro-DiasMC, SantosH (1990) Effects of ethanol on Saccharomyces cerevisiae as monitored by in vivo 31P and 31C nuclear magnetic resonance. Arch Microbiol 153: 384–391
MillarDG, Griffiths-SmithK, AlgarE, ScopesRK (1982) Activity and stability of glycolytic enzymes in the presence of ethanol. Biotechnol Lett 4: 601–606
NegeleinE, NollF (1984) d-Glycerate 1,3-bisphosphate. In: BergmeyerHU, BergmeyerJ, GrasslH (eds) Methods of enzymatic analysis, vol 6. Verlag Chemie, Weinheim pp 542–546
OsmanYA, IngramLO (1985) Mechanism of ethanol inhibition of fermentation in Zymomonas mobilis CP4. J Bacteriol 164: 173–180
OsmanYA, ConwayT, BonettiSJ, IngramLO (1987) Glycolytic flux in Zymomonas mobilis: enzyme and metabolite levels during batch fermentation. J Bacteriol 169: 3726–3736
RogersPL, LeeKJ, SkotnickiML, TribeDE (1982) Ethanol production by Zymomonas mobilis. Adv Biochem Engin 23: 37–84
RottenbergH (1979) The measurement of membrane potential and delta pH in cells, organelles, and vesicles. In: FleischerS, PackerL (eds) Methods in enzymology, vol 55. Academic Press, New York, pp 547–569
ScopesRK, Griffiths-SmithK (1984) Use of differential dye-ligand chromatography with affinity elution for enzyme purification: 6-phosphogluconate dehydratase from Zymomonas mobilis. Anal Biochem 136: 530–534
ScopesRK, TestolinV, StoterA, Griffiths-SmithK, AlgarEM (1985) Simultaneous purification and characterization of glucokinase, fructokinase and glucose-6-phosphate dehydrogenase from Zymomonas mobilis. Biochem J 228: 627–634
SwingsJ, DeLeyJ (1977) The biology of Zymomonas. Bacteriol Rev 41: 1–46
UgurbilK, ShulmanRG, BrownTR (1979) High-resolution 31P and 13C nuclear magnetic resonance studies of Escherichia coli cells in vivo. In: ShulmanRG (ed) Biological applications of magnetic resonance. Academic Press, New York, pp 537–589
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Strohhäcker, J., de Graaf, A.A., Schoberth, S.M. et al. 31P Nuclear magnetic resonance studies of ethanol inhibition in Zymomonas mobilis . Arch. Microbiol. 159, 484–490 (1993). https://doi.org/10.1007/BF00288598
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DOI: https://doi.org/10.1007/BF00288598