Metabolic flux rearrangement in the amino acid metabolism reduces ammonia stress in the α1-antitrypsin producing human AGE1.HN cell line

This study focused on metabolic changes in the neuronal human cell line AGE1.HN upon increased ammonia stress. Batch cultivations of α1-antitrypsin (A1AT) producing AGE1.HN cells were carried out in media with initial ammonia concentrations ranging from 0 mM to 5 mM. Growth, A1AT production, metabolite dynamics and finally metabolic fluxes calculated by metabolite balancing were compared. Growth and A1AT production decreased with increasing ammonia concentration. The maximum A1AT concentration decreased from 0.63 g/l to 0.51 g/l. Central energy metabolism remained relatively unaffected exhibiting only slightly increased glycolytic flux at high initial ammonia concentration in the medium. However, the amino acid metabolism was significantly changed. Fluxes through transaminases involved in amino acid degradation were reduced concurrently with a reduced uptake of amino acids. On the other hand fluxes through transaminases working in the direction of amino acid synthesis, i.e., alanine and phosphoserine, were increased leading to increased storage of excess nitrogen in extracellular alanine and serine. Glutamate dehydrogenase flux was reversed increasingly fixing free ammonia with increasing ammonia concentration. Urea production additionally observed was associated with arginine uptake by the cells and did not increase at high ammonia stress. It was therefore not used as nitrogen sink to remove excess ammonia. The results indicate that the AGE1.HN cell line can adapt to ammonia concentrations usually present during the cultivation process to a large extent by changing metabolism but with slightly reduced A1AT production and growth.

► The human A1AT producing cell line AGE1.HN can cope with high ammonia stress.
► Ammonia stress is reduced by modified metabolic activities.
► Metabolic flux analysis sheds light on the detailed metabolic changes.
► Ammonia addition causes increased secretion of certain amino acids, e.g., alanine.
► Reduced uptake of metabolized amino acids, e.g., leucine, decreases ammonia stress.