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Tailoring Escherichia coli for the L-rhamnose PBAD promoter-based production of membrane and secretory proteins

Hjelm, A., Karyolaimos, A., Zhang, Z., Rujas, E., Vikström, D., Slotboom, D. J. & de Gier, J-W., Apr-2017, In : ACS Synthetic Biology. 6, 6, p. 985-994 10 p., acssynbio.6b00321.

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  • Tailoring Escherichia coli for the L-rhamnose PBAD promoter-based production of membrane and secretory proteins

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DOI

  • Anna Hjelm
  • Alexandros Karyolaimos
  • Zhe Zhang
  • Edurne Rujas
  • David Vikström
  • Dirk Jan Slotboom
  • Jan-Willem de Gier

Membrane and secretory protein production in Escherichia coli requires precisely controlled production rates to avoid the deleterious saturation of their biogenesis pathways. Based on this requirement, the E. coli L-rhamnose PBAD promoter (PrhaBAD) is often used for membrane and secretory protein production since PrhaBAD is thought to regulate protein production rates in an L-rhamnose concentration dependent manner. By monitoring protein production in real-time in E. coli wild-type and an L-rhamnose catabolism deficient mutant, we demonstrate that the L-rhamnose concentration-dependent tunability of PrhaBAD-mediated protein production is actually due to L-rhamnose consumption rather than regulating production rates. Using this information, a RhaT-mediated L-rhamnose transport and L-rhamnose catabolism deficient double mutant was constructed. We show that this mutant enables to regulate PrhaBAD-based protein production rates in an L-rhamnose concentration dependent manner and that this is critical to optimize membrane and secretory protein production yields. The high precision of protein production rates provided by the PrhaBAD promoter in an L-rhamnose transport and catabolism deficient background could also benefit other applications in synthetic biology.

Original languageEnglish
Article numberacssynbio.6b00321
Pages (from-to)985-994
Number of pages10
JournalACS Synthetic Biology
Volume6
Issue number6
Early online date22-Feb-2017
Publication statusPublished - Apr-2017

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