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Growth characteristics of Saccharomyces cerevisiae S288C in changing environmental conditions: auxo-accelerostat study

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Abstract

The effect of individual environmental conditions (pH, pO2, temperature, salinity, concentration of ethanol, propanol, tryptone and yeast extract) on the specific growth rate as well as ethanol and glycerol production rate of Saccharomyces cerevisiae S288C was mapped during the fermentative growth in aerobic auxo-accelerostat cultures. The obtained steady-state values of the glycerol to ethanol formation ratio (0.1 mol mol−1) corresponding to those predicted from the stoichiometric model of fermentative yeast growth showed that the complete repression of respiration was obtained in auxostat culture and that the model is suitable for calculation of Y ATP and Q ATP values for the aerobic fermentative growth. Smooth decrease in the culture pH and dissolved oxygen concentration (pO2) down to the critical values of 2.3 and 0.8%, respectively, resulted in decrease in growth yield (Y ATP) and specific growth rate, however the specific ATP production rate (Q ATP) stayed almost constant. Increase in the concentration of biomass (>0.8 g dwt l−1), propanol (>2 g l−1) or NaCl (>15 g l−1) lead at first to the decrease in the specific growth rate and Q ATP, while Y ATP was affected only at higher concentrations. The observed decrease in Q ATP was caused by indirect rather than direct inhibition of glycolysis. The increase in tryptone concentration resulted in an increase in the specific growth rate from 0.44 to 0.62 h−1 and Y ATP from 12.5 to 18.5 mol ATP g dwt−1. This study demonstrates that the auxo-accelerostat method, besides being an efficient tool for obtaining the culture characteristics, provides also decent conditions for the experiments elucidating the control mechanisms of cell growth.

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Acknowledgements

The financial support for this research provided by Estonian Ministry of Education and Research (targeted funding projects 0222601Bs03 and 0142497s03) and EU Structural Funds is gratefully acknowledged.

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Authors and Affiliations

  1. National Institute of Chemical Physics and Biophysics, Akadeemia tee 23, 12618, Tallinn, Estonia

    Kaja Kasemets

  2. Tallinn University of Technology, Ehitajate tee 5, 19086, Tallinn, Estonia

    Ildar Nisamedtinov, Tiiu-Maie Laht, Kristo Abner & Toomas Paalme

  3. Competence Centre of Food and Fermentation Technologies, Akadeemia tee 15, 12618, Tallinn, Estonia

    Ildar Nisamedtinov, Kristo Abner & Toomas Paalme

Authors
  1. Kaja Kasemets
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  2. Ildar Nisamedtinov
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  3. Tiiu-Maie Laht
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  4. Kristo Abner
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  5. Toomas Paalme
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Corresponding author

Correspondence to Toomas Paalme.

Appendices

Appendix 1

Stoichiometric model of anaerobic growth of S. cerevisiae

The set of key reactions of central metabolism supporting the fermentative growth of Saccharomyces cerevisiae is shown in Fig. 1. Assuming that the concentration of central metabolites remains constant in the cells during exponential growth of the cells, the mass balance equations for each key intermediate can be written:

1. Glucose-6-P:

rGLC = PGI +  ZWF1 +  o 1

2. 6-P-glyconolactone:

ZWF1 = pgl

3. 6-P-glyconate:

pgl =  GND1,2

4. Ribulose-5-P:

GND1,2 =  RKI1 + RPE1

5. Xylulose-5-P:

RPE1 = TLK1,2a +  TLK1,2b

6. Ribose-5-P:

RKI1 =  TLK1,2a +  o 3

7. Seduheptulose-7-P:

TLK1,2a  = TAL1

8. Erythrose-4-P:

TAL1 = TLK1,2b +  o 4

9. Fructose-6-P:

PGI +  TLK1,2b  + TAL1 = PFK1,2 +  o 2

10. Fructose-1,6-P:

PFK1,2 =  FBA1

11. Dihydroxyacetone-P:

FBA1 = TPI1 +  r GLR

12. Glyceraldehyde-3-P:

TLK1,2a +  TLK1,2b  + FBA1 + TPI1 = TAL1 + TDH1,2,3 +  o 5

13. 1,3-Diphosphoglycerate:

TDH1,2,3 = PGK

14. 3-Phosphoglycerate:

PGK =  GPM +  o 6

15. 2-Phosphoglycerate:

GPM =  ENO

16. Phosphoenolpyruvate:

ENO =  PYK1 +  o 7

17. NADPH:

ZWF1 +  GND1,2 + IDP1 + ALD6 =  o NADP

18. 2-Ketoglutarate:

IDP1 =  o 11

19. Oxaloacetate m :

OAC =  CIT

20. Acetyl-CoA m :

CIT =  YAT

21. Citrate:

CIT =  IDP1

22. Oxaloacetate c :

OAC +  o 10 = PYC

23. Pyruvate:

PYK1 =  o 8 + PDC +  PYC

24. Acetalaldehyde

PDC =  r ETH + ALD6

25. Acetyl-CoA c :

ALD6 = YAT +  o 9

26. NADH:

TDH1,2,3 =  r ETH +  r GLR +  o NADH

27. ATP:

q ATP = −r GLC − PFK1,2 +  PGK +  PYK1−PYC− 2* ALD6

The equation system consists of 27 formulas (metabolites) and 41 variables (relative net fluxes). The 11 relative fluxes o i (mmol g dwt−1) represent the anabolic demands of corresponding key intermediates (Fig. 1), and oNADH, oNADPH the anabolic demands of corresponding co-factors required for biomass synthesis. If the macromolecular composition of the cells is known, the values of anabolic demands can be determined from the composed stoichiometric matrix, similar to that presented by Cortassa et al. (1995). The three fluxes r GLC, r GLR and r ETH are experimentally measured (r i  = Q i /μ, Formula 2 and 3). As altogether 16 variables can be calculated or measured the equation system becomes over determined (2 determined variables more than the degrees of freedom).

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Kasemets, K., Nisamedtinov, I., Laht, TM. et al. Growth characteristics of Saccharomyces cerevisiae S288C in changing environmental conditions: auxo-accelerostat study. Antonie van Leeuwenhoek 92, 109–128 (2007). https://doi.org/10.1007/s10482-007-9141-y

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