Recombinant Protein Production with Escherichia coli in Glucose and Glycerol Limited Chemostats

Anca Manuela Mitchell, Valentina Gogulancea, Wendy Smith, Anil Wipat, Irina Dana Ofiţeru

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Recently, there has been a resurgence of interest in continuous bioprocessing as a cost-optimised production strategy, driven by a rising global requirement for recombinant proteins used as biological drugs. This strategy could provide several benefits over traditional batch processing, including smaller bioreactors, smaller facilities, and overall reduced plant footprints and investment costs. Continuous processes may also offer improved product quality and minimise heterogeneity, both in the culture and in the product. In this paper, a model protein, green fluorescent protein (GFP) mut3*, was used to test the recombinant protein expression in an Escherichia coli strain with industrial relevance grown in chemostat. An important factor in enabling stable productivity in continuous cultures is the carbon source. We have studied the viability and heterogeneity of the chemostat cultures using a chemically defined medium based on glucose or glycerol as the single carbon source. As a by-product of biodiesel production, glycerol is expected to become a sustainable alternative substrate to glucose. We have found that although glycerol gives a higher cell density, it also generates higher heterogeneity in the culture and a less stable recombinant protein production. We suggest that manipulating the balance between different subpopulations to increase the proportion of productive cells may be a possible solution for making glycerol a successful alternative to glucose.
Original languageEnglish
Pages (from-to)239-254
Number of pages16
JournalApplied Microbiology
Issue number2
Early online date16 Jul 2021
Publication statusPublished online - 16 Jul 2021


  • Escherichia coli
  • recombinant protein production
  • chemostat
  • carbon source
  • flow cytometry


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