Unraveling the metabolism of HEK-293 cells using lactate isotopomer analysis |
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Authors: | Olivier Henry Mario Jolicoeur Amine Kamen |
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Institution: | (1) Chemical Engineering Department, ?cole Polytechnique de Montr?al, C.P. 6079, Succ. Centre-ville, Montr?al, QC, H3C 3A7, Canada;(2) National Research Council Canada, Biotechnology Research Institute, 6100 Royalmount Avenue, Montr?al, QC, H4P 2R2, Canada |
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Abstract: | HEK-293 is the most extensively used human cell line for the production of viral vectors and is gaining increasing attention
for the production of recombinant proteins by transient transfection. To further improve the metabolic characterization of
this cell line, we have performed cultures using 13C-labeled substrates and measured the resulting mass isotopomer distributions in lactate by LC/MS. Simultaneous metabolite
and isotopomer balancing allowed improvement and validation of the metabolic model and quantification of key intracellular
pathways. We have determined the amounts of glucose carbon channeled through the PPP, incorporated into the TCA cycle for
energy production and lipids biosynthesis, as well as the cytosolic and mitochondrial malic enzyme fluxes. Our analysis also
revealed that glutamine did not significantly contribute to lactate formation. An improved and quantitative understanding
of the central carbon metabolism is greatly needed to pursue the rational development of engineering approaches at both the
cellular and process levels. |
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Keywords: | |
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