Activity of glutamate dehydrogenase is increased in ammonia-stressed hybridoma cells.
The effect of added ammonia on the intracellular fluxes in hybridoma cells was investigated by metabolic-flux balancing techniques. It was found that, in ammonia-stressed hybridoma cells, the glutamate-dehydrogenase flux is in the reverse direction compared to control cells. This demonstrates that hybridoma cells are able to prevent the accumulation of ammonia by converting ammonia and alpha-ketoglutarate into glutamate. The additional glutamate that is produced by this flux, as compared to the control culture, is converted by the reactions catalyzed by alanine aminotransferase (45% of the extra glutamate) and aspartate aminotransferase (37%), and a small amount is used for the biosynthesis of proline (6%). The remaining 12% of the extra glutamate is secreted into the culture medium. The data suggest that glutamate dehydrogenase is a potential target for metabolic engineering to prevent ammonia accumulation in high-cell-density culture.
Bonarius
H P
HP
Department PRP-Biotechnology, Hoffmann-La Roche Ltd., Basel, Switzerland.Bona@Novo.dk
Houtman
J H
JH
de Gooijer
C D
CD
Tramper
J
J
Schmid
G
G
D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002244:Carbon / Q000378:metabolism; D002455:Cell Division; D002470:Cell Survival; D002478:Cells, Cultured; D005969:Glutamate Dehydrogenase / Q000187:drug effects / Q000378:metabolism*; D006825:Hybridomas / Q000187:drug effects* / Q000201:enzymology*; D008322:Mammals; D009584:Nitrogen / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism / Q000494:pharmacology*
D000596:Amino Acids; D000644:Quaternary Ammonium Compounds; D002244:Carbon; D005969:Glutamate Dehydrogenase; D009584:Nitrogen