92
Chapter 3
Problem 3.6 Comparison between two strains of
S. cerevisiae.
Nissen
et al.
(2001) studied anaerobic cultivation of three different strains of
S. cerevisiae,
TNI, TN26
and TN21 on a defined medium with glucose as carbon and energy source. TNI is the “standard” haploid
strain used in a number of studies by the authors. TN21 is a transformant of TNI which expresses the
transhydrogenase gene
tdh
from
Azotobacter vinlandii
to a high level, 4.53 units mg'1. TN26 is also a
transformant of TNI with the same multicopy expression vector as in TNI, but without ligation of the
tdh
gene. TN26 is consequently genetically identical to TN21, except that it cannot express
transhydrogenase. It is therefore unable to equilibrate the reversible reaction between NADPH/NAD+ and
NADH/NADP*.
The table below shows product yields from steady state, glucose limited continuous cultivations of the
three strains. Note the high accuracy of the data, and also that TN26 behaves exactly as TNI. There are,
however, small but significant differences between TN21 and TN26. All data are given as C-mole (C-
mole glucose)1.
Product
TNI
TN26
TN21
Ethanol
0.494 ±0.012
0.493 ±0.011
0.451 ±0.011
Glycerol
0.091 ± 0.001
0.093 ±0.001
0.118 ±0.001
CQ.
0.275 ± 0.007
0.271 ±0.007
0.262 ±0.007
Biomass
0.111 ±0.002
0.112 ±0.002
0.102 ±0.002
Succinate
0.005 ±0.0003
0.005 ± 0.0003
0.006 ± 0.0003
Pyruvate
0.005 ± 0.0003
0.005 ± 0.0003
0.001 ± 0.0003
Acetate
0.005 ± 0.0003
0.005 ± 0.0003
0.007 ± 0.0003
2-Oxaloacetate
0.002 ±0.0003
0.007 ±0.0003
0.045 ± 0.0003
Total
0.988 ± 0.022
0.991 ±0.021
0.992 ±0.021
a.
Check the redox balance for TN26 and TN21.
b.
Assume that the missing carbon is ethanol. Repeat the calculation of a with this assumption.
c.
Discuss qualitatively the difference between the product profiles for TN26 and TN21. What
could be the reason for the higher yield of 2-oxaloacetate (=a ketoglutarate)? The dominant
cofactor for conversion of 2-oxoglutarate to glutamic acid is NAPDH in all three strains.
REFERENCES
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Paracoccus pantotrophus
under dynamic substrate supply.
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55,773-782.
Andersen, M.Y., Pedersen, N.
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bioreactor near the critical dilution rate using a productostat.
J. BiotechnoL,
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Bijkerk, A.H.E, and Hall, RJ. (1977). A mechanistic model of the aerobic growth of
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Christensen .L.H., Schulze, U., Nielsen,J., and Villadsen, J. (1995). Acoustic gas analysis for fast and precise monitoring of
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Cook, A. H. (1958).
The Chemistry and Biology of Yeasts,
Academic Press, New York.
Duboc, P., von Stockar, U. (1998). Systematic errors in data evaluation due to ethanol stripping and water vaporization.
BiotechnoL Bioeng.
58,428-439
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