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      Online measurement of the respiratory activity in shake flasks enables the identification of cultivation phases and patterns indicating recombinant protein production in various Escherichia coli host strains.

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          Abstract

          Escherichia coli is commonly used for recombinant protein production with many available host strains. Screening experiments are often performed in batch mode using shake flasks and evaluating only the final product concentration. This conventional approach carries the risk of missing the best strain due to limited monitoring capabilities. Thus, this study focuses on investigating the general suitability of online respiration measurement for selecting expression hosts for heterologous protein production. The oxygen transfer rate (OTR) for different T7-RNA polymerase-dependent Escherichia coli expression strains was compared under inducing and noninducing conditions. As model enzymes, a lipase A from Bacillus subtilis (BSLA) and a 3-hydroxybutyryl-CoA dehydrogenase from Thermus thermophilus (HBD) were chosen. Four strains were compared during expression of both enzymes in autoinduction medium. Additionally, four strains were compared during expression of the BSLA with IPTG induction. It was found that the metabolic burden during recombinant protein production induces a phase of constant OTR, while undisturbed cell growth with no or little product formation is indicated by an exponential increase. This pattern is independent of the host strain, expressed enzyme, and induction method. Furthermore, the OTR gives information about carbon source consumption, biomass formation, and the transition from production to noninduced second growth phase, thereby ensuring a fair comparison of different strains. In conclusion, online monitoring of the respiration activity is suited to qualitatively identify, if a recombinant protein is produced by a strain or not. Furthermore, laborious offline sampling is avoided. Thus, the technique is easier and faster compared to conventional approaches. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:315-327, 2018.

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          Author and article information

          Journal
          Biotechnol Prog
          Biotechnology progress
          Wiley
          1520-6033
          1520-6033
          March 2018
          : 34
          : 2
          Affiliations
          [1 ] AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstr. 51, Aachen D-52074, Germany.
          [2 ] Bioeconomy Science Center (BioSC), Jülich, Germany.
          [3 ] Institute of Molecular Enzyme Technology, Heinrich-Heine-University Düsseldorf, Forschungszentrum Jülich, Jülich D-52426, Germany.
          [4 ] Institute of Bio- and Geosciences IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich D-52426, Germany.
          [5 ] Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan.
          Article
          10.1002/btpr.2600
          29314728
          5b2c795a-a43c-4d3b-b2af-84d7dd099789
          © 2018 American Institute of Chemical Engineers.
          History

          IPTG,T7-based E. coli host strains,autoinduction,online monitoring,recombinant protein production

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