Recombinant Fab expression and secretion in Escherichia coli continuous culture at medium cell densities: Influence of temperature

Escarlata Rodríguez-Carmona, Olivia Cano-Garrido, Martin Dragosits, Michael Maurer, Alexander Mader, Renate Kunert, Diethard Mattanovich, Antonio Villaverde, Felícitas Vázquez

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18 Citations (Scopus)

Abstract

Production of recombinant antibody fragments (Fabs) in Escherichia coli has gained interest because of the recognised advantages of this expression system and because Fabs do not require glycosylation. However, more comprehensive studies on the factors that influence expression conditions and product yield are still required for full process development. In this work, the effect of growth temperature on the periplasmatic expression of the 3H6 Fab in E. coli was studied in carbon-limited continuous cultures operated at medium cell densities. Three different temperatures were assayed, namely 37, 33 and 30 °C. Results showed that biomass yield was not affected within this temperature range whilst product yield increased as temperature decreased. Periplasmic Fab secretion corresponded to 30% of the produced Fab protein and its efficiency was irrespective of the process temperature. Moreover, considerable product leakage to the culture supernatant was detected in all cases, ranging from about 40% at 37 °C to almost 70% at 30 °C. Besides, plasmid loss was observed along process time indicating a selective pressure against plasmid-bearing cells. This study supports the potential of continuous cultivations of E. coli at medium cell densities under well controlled conditions as a tool for characterising the impact of environmental parameters and cell physiology under protein production conditions. © 2011 Elsevier Ltd. All rights reserved.
Original languageEnglish
Pages (from-to)446-452
JournalProcess Biochemistry
Volume47
Issue number3
DOIs
Publication statusPublished - 1 Mar 2012

Keywords

  • Continuous culture
  • E. coli
  • Fab production
  • Process design
  • Process monitoring
  • Temperature influence

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    Rodríguez-Carmona, E., Cano-Garrido, O., Dragosits, M., Maurer, M., Mader, A., Kunert, R., Mattanovich, D., Villaverde, A., & Vázquez, F. (2012). Recombinant Fab expression and secretion in Escherichia coli continuous culture at medium cell densities: Influence of temperature. Process Biochemistry, 47(3), 446-452. https://doi.org/10.1016/j.procbio.2011.11.024