Details
Original language | English |
---|---|
Article number | 50 |
Journal | BMC Systems Biology |
Volume | 8 |
Issue number | 1 |
Publication status | Published - 28 Apr 2014 |
Externally published | Yes |
Abstract
Background: Mapping the intracellular fluxes for established mammalian cell lines becomes increasingly important for scientific and economic reasons. However, this is being hampered by the high complexity of metabolic networks, particularly concerning compartmentation.Results: Intracellular fluxes of the CHO-K1 cell line central carbon metabolism were successfully determined for a complex network using non-stationary 13C metabolic flux analysis. Mass isotopomers of extracellular metabolites were determined using [U-13C6] glucose as labeled substrate. Metabolic compartmentation and extracellular transport reversibility proved essential to successfully reproduce the dynamics of the labeling patterns. Alanine and pyruvate reversibility changed dynamically even if their net production fluxes remained constant. Cataplerotic fluxes of cytosolic phosphoenolpyruvate carboxykinase and mitochondrial malic enzyme and pyruvate carboxylase were successfully determined. Glycolytic pyruvate channeling to lactate was modeled by including a separate pyruvate pool. In the exponential growth phase, alanine, glycine and glutamate were excreted, and glutamine, aspartate, asparagine and serine were taken up; however, all these amino acids except asparagine were exchanged reversibly with the media. High fluxes were determined in the pentose phosphate pathway and the TCA cycle. The latter was fueled mainly by glucose but also by amino acid catabolism.Conclusions: The CHO-K1 central metabolism in controlled batch culture proves to be robust. It has the main purpose to ensure fast growth on a mixture of substrates and also to mitigate oxidative stress. It achieves this by using compartmentation to control NADPH and NADH availability and by simultaneous synthesis and catabolism of amino acids.
Keywords
- Chinese hamster ovary cells, CHO, Compartmentation, Mammalian cell culture, Mammalian metabolism, Metabolic flux analysis, Metabolic transport, Mitochondria, Reversibility
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Structural Biology
- Mathematics(all)
- Modelling and Simulation
- Biochemistry, Genetics and Molecular Biology(all)
- Molecular Biology
- Computer Science(all)
- Computer Science Applications
- Mathematics(all)
- Applied Mathematics
Cite this
- Standard
- Harvard
- Apa
- Vancouver
- BibTeX
- RIS
In: BMC Systems Biology, Vol. 8, No. 1, 50, 28.04.2014.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Non-stationary 13C metabolic flux analysis of Chinese hamster ovary cells in batch culture using extracellular labeling highlights metabolic reversibility and compartmentation
AU - Nicolae, Averina
AU - Wahrheit, Judith
AU - Bahnemann, Janina
AU - Zeng, An Ping
AU - Heinzle, Elmar
N1 - Funding information: We thank the Institute of Cell Culture Technology (University Bielefeld, Germany) for supplying the CHO K1 cells and the BMBF (German Federal Ministry of Education and Research) project SysCompart (project ID 031555D), part of the Systems Biology program, New Methods in Systems Biology (SysTec), for funding.
PY - 2014/4/28
Y1 - 2014/4/28
N2 - Background: Mapping the intracellular fluxes for established mammalian cell lines becomes increasingly important for scientific and economic reasons. However, this is being hampered by the high complexity of metabolic networks, particularly concerning compartmentation.Results: Intracellular fluxes of the CHO-K1 cell line central carbon metabolism were successfully determined for a complex network using non-stationary 13C metabolic flux analysis. Mass isotopomers of extracellular metabolites were determined using [U-13C6] glucose as labeled substrate. Metabolic compartmentation and extracellular transport reversibility proved essential to successfully reproduce the dynamics of the labeling patterns. Alanine and pyruvate reversibility changed dynamically even if their net production fluxes remained constant. Cataplerotic fluxes of cytosolic phosphoenolpyruvate carboxykinase and mitochondrial malic enzyme and pyruvate carboxylase were successfully determined. Glycolytic pyruvate channeling to lactate was modeled by including a separate pyruvate pool. In the exponential growth phase, alanine, glycine and glutamate were excreted, and glutamine, aspartate, asparagine and serine were taken up; however, all these amino acids except asparagine were exchanged reversibly with the media. High fluxes were determined in the pentose phosphate pathway and the TCA cycle. The latter was fueled mainly by glucose but also by amino acid catabolism.Conclusions: The CHO-K1 central metabolism in controlled batch culture proves to be robust. It has the main purpose to ensure fast growth on a mixture of substrates and also to mitigate oxidative stress. It achieves this by using compartmentation to control NADPH and NADH availability and by simultaneous synthesis and catabolism of amino acids.
AB - Background: Mapping the intracellular fluxes for established mammalian cell lines becomes increasingly important for scientific and economic reasons. However, this is being hampered by the high complexity of metabolic networks, particularly concerning compartmentation.Results: Intracellular fluxes of the CHO-K1 cell line central carbon metabolism were successfully determined for a complex network using non-stationary 13C metabolic flux analysis. Mass isotopomers of extracellular metabolites were determined using [U-13C6] glucose as labeled substrate. Metabolic compartmentation and extracellular transport reversibility proved essential to successfully reproduce the dynamics of the labeling patterns. Alanine and pyruvate reversibility changed dynamically even if their net production fluxes remained constant. Cataplerotic fluxes of cytosolic phosphoenolpyruvate carboxykinase and mitochondrial malic enzyme and pyruvate carboxylase were successfully determined. Glycolytic pyruvate channeling to lactate was modeled by including a separate pyruvate pool. In the exponential growth phase, alanine, glycine and glutamate were excreted, and glutamine, aspartate, asparagine and serine were taken up; however, all these amino acids except asparagine were exchanged reversibly with the media. High fluxes were determined in the pentose phosphate pathway and the TCA cycle. The latter was fueled mainly by glucose but also by amino acid catabolism.Conclusions: The CHO-K1 central metabolism in controlled batch culture proves to be robust. It has the main purpose to ensure fast growth on a mixture of substrates and also to mitigate oxidative stress. It achieves this by using compartmentation to control NADPH and NADH availability and by simultaneous synthesis and catabolism of amino acids.
KW - Chinese hamster ovary cells
KW - CHO
KW - Compartmentation
KW - Mammalian cell culture
KW - Mammalian metabolism
KW - Metabolic flux analysis
KW - Metabolic transport
KW - Mitochondria
KW - Reversibility
UR - http://www.scopus.com/inward/record.url?scp=84899952895&partnerID=8YFLogxK
U2 - 10.1186/1752-0509-8-50
DO - 10.1186/1752-0509-8-50
M3 - Article
C2 - 24773761
AN - SCOPUS:84899952895
VL - 8
JO - BMC Systems Biology
JF - BMC Systems Biology
SN - 1752-0509
IS - 1
M1 - 50
ER -