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Enhancement of extracellular pullulanase production from recombinant Escherichia coli by combined strategy involving auto-induction and temperature control

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Abstract

Pullulanase was extracellularly produced with an engineered Escherichia coli with a combined strategy. When auto-induction instead of isopropyl β-d-1-thiogalactopyranoside (IPTG) induction method was implemented, we observed increased extracellular activity (4.2 U ml−1) and cell biomass (7.95 g DCW l−1). Subsequent investigation of temperature effect on fermentation showed cultivation performed at 25 °C presented the highest extracellular titer and cell biomass. In order to reduce the extended production period, we developed a two-stage temperature control strategy. Its application not only reduced the production period from 72 to 36 h, but also further enhanced the yield of extracellular pullulanase. Finally, with a view to releasing more intracellular pullulanase, we altered cell membrane permeability with various medium additives. As a result, extracellular titer was elevated to 68.23 U ml−1, nearly 35-fold higher than that with IPTG induction method. The combined strategy developed here may be useful for the production of other extracellular proteins by recombinant E. coli.

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Acknowledgments

Financial supports from the Hi-Tech Research and Development Program of China (863 Program) (no. 2012AA022207), the National Key Basic Research and Development Program of China (973 Program) (no. 2011CB710800 and 2009CB724706), the Program of Introducing Talents of Discipline to Universities (111 Project) (111-2-06), and the High-end Foreign Experts Recruitment Program (GDW20123200113) are gratefully appreciated.

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Authors and Affiliations

  1. Key Laboratory of Industrial Biotechnology, Ministry of Education and School of Biotechnology, Jiangnan University, 1800 Lihu Ave., Wuxi, 214122, Jiangsu Province, China

    Wen-Bo Chen, Yao Nie & Yan Xu

  2. Department of Molecular Biology and Biochemistry, Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, NJ, 08854, USA

    Rong Xiao

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  1. Wen-Bo Chen
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  2. Yao Nie
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  3. Yan Xu
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Correspondence to Yao Nie or Yan Xu.

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Chen, WB., Nie, Y., Xu, Y. et al. Enhancement of extracellular pullulanase production from recombinant Escherichia coli by combined strategy involving auto-induction and temperature control. Bioprocess Biosyst Eng 37, 601–608 (2014). https://doi.org/10.1007/s00449-013-1026-z

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  • DOI: https://doi.org/10.1007/s00449-013-1026-z

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