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Metabolic characteristics of recombinant Chinese hamster ovary cells expressing glutamine synthetase in presence and absence of glutamine

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Abstract

To elucidate the metabolic characteristics of recombinant CHO cells expressing glutamine synthetase (GS) in the medium with or without glutamine, the concentrations of extra- and intracellular metabolites and the activities of key metabolic enzymes involved in glutamine metabolism pathway were determined. In the absence of glutamine, glutamate was utilized for glutamine synthesis, while the production of ammonia was greatly decreased. In addition, the expression of recombinant protein was increased by 18%. Interestingly, the intracellular glutamine maintained almost constant, independent of the presence of glutamine or not. Activities of glutamate-oxaloacetate aminotransferase (GOT), glutamate-pyruvate aminotransferase (GPT), and glutamate dehydrogenase (GDH) increased in the absence of glutamine. On the other hand, intracellular isocitrate and the activities of its downstream isocitrate dehydrogenase in the TCA cycle increased also. In combination with these two factors, a 8-fold increase in the intracellular α-ketoglutarate was observed in the culture of CHO-GS cells in the medium without glutamine.

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Abbreviations

CHO-GS:

Recombinant Chinese hamster ovary cell bearing the exogenous glutamine synthetase gene

GS:

Glutamine synthetase

GOT:

Glutamate-oxaloacetate aminotransferase

GPT:

Glutamate-pyruvate aminotransferase

GDH:

Glutamate dehydrogenase

PAG:

Phosphate-activated glutaminase

C X :

Viable cell density

α-KG:

α-Ketoglutarate

Fum:

Fumarate

Cit:

Citrate

Icit:

Isocitrate

Mal:

Malate

MSX:

Methionine sulphoximine

Q P :

Specific production rate of metabolite

Q S :

Specific consumption rate of substrate

Y P/S :

Apparent yield coefficient of product to substrate

0:

Seeding time

t:

Harvest time

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Acknowledgements

This work was supported by the grants from the National High-Tech Research and Development Programs of China (No. 2004AA2Z3792).

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  1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China

    Fang Zhang, Xiangming Sun, Xiaoping Yi & Yuanxing Zhang

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  1. Fang Zhang
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  2. Xiangming Sun
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  3. Xiaoping Yi
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  4. Yuanxing Zhang
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Correspondence to Yuanxing Zhang.

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Zhang, F., Sun, X., Yi, X. et al. Metabolic characteristics of recombinant Chinese hamster ovary cells expressing glutamine synthetase in presence and absence of glutamine. Cytotechnology 51, 21–28 (2006). https://doi.org/10.1007/s10616-006-9010-y

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  • DOI: https://doi.org/10.1007/s10616-006-9010-y

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