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Metabolic and bioprocess engineering of production cell lines for recombinant protein production

Bielefelder Schriften zur Zellkulturtechnik, Bd. 4

Betina da Silva Ribeiro

ISBN 978-3-8325-3123-2
160 Seiten, Erscheinungsjahr: 2012
Preis: 53.50 €
Dissolved carbon dioxide (pCO2) has been identified as an important process parameter affecting cell growth, productivity and product quality (e.g. glycosylation) of recombinant proteins when exceeding critical levels, occurring especially in industrial large-scale cell culture processes due to the increased hydrostatic pressure. As CO2 can easily pass the cellular membrane and thereby influence intracellular pH (pHi), important cellular processes (e.g. cell cycle regulation, enzymes of TCA cycle) are directly influenced by pCO2 and dependent bicarbonate concentration. Consequently, process control strategies attend to keep pCO2 within physiological range.

In a metabolic engineering approach an antibody producing CHO cell line stably expressing human carbonic anhydrase (hCAII), the enzyme that catalyzes the equilibrium of CO2 in aqueous solutions, was generated and used to characterize CO2 effects in simulated CO2 acid load and high CO2 levels as they occur in large scale mammalian cell culture. The cell line expressing active hCAII showed more rapid initial re-alkalinization of cytoplasm after induced CO2 acid load. Results also suggest that cellular pHi fine tuning was performed by the Cl-/HCO3- exchanger (AE) and Na-dependent Cl-/HCO3- exchanger (NCBE) instead of the Na+/H+ exchanger (NHE1).


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Keywords:
  • carbon dioxide
  • metabolic engineering
  • human carbonic anhydrase
  • intracellular pH
  • recombinant proteins

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