INDUCTION OF A METABOLIC SWITCH IN INSECT CELLS BY SUBSTRATE LIMITED FED-BATCH CULTURES

Insect cell metabolism was studied in substrate-limited fed batch cult ures of Spodoptera frugiperda (Sf-9) cells. Results from a glucose-lim ited culture, a glutamine-limited culture, a culture limited in both g lucose and glutamine and a batch culture were compared. A stringent re lation between glucose excess and alanine formation was found. In cont rast, glucose limitation induced ammonium formation, while, at the sam e time, alanine formation was completely suppressed. Simultaneous gluc ose and glutamine limitation suppressed both alanine and ammonium form ation. Although the metabolism was influenced by substrate limitation, the specific growth rate was similar in all cultures. Alanine formati on must involve incorporation of free ammonium, if ammonium formation is mediated by glutaminase and glutamate dehydrogenase, as our data su ggest. On the basis of the results, two possible pathways for the form ation of alanine in the intermediary metabolism in insect cells are su ggested. The cellular yield on glucose was increased 6.6 times during glucose limitation, independently of the cellular yield on glutamine, which was increased 50-100 times during glutamine limitation. The resu lts indicate that alanine overflow metabolism is energetically wastefu l and that glutamine is a dispensable amino acid for cultured Sf-9 cel ls. Preliminary data confirm that glutamine can be synthesised by the cells themselves in amounts sufficient to support growth.