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Applied Microbiology and Biotechnology

, Volume 72, Issue 3, pp 529–536 | Cite as

Characterization of the T7 promoter system for expressing penicillin acylase in Escherichia coli

  • Yali Xu
  • Stefan Rosenkranz
  • Chiao-Ling Weng
  • Jeno M. Scharer
  • Murray Moo-Young
  • C. Perry ChouEmail author
Applied Genetics and Molecular Biotechnology

Abstract

The pac gene encoding penicillin acylase (PAC) was overexpressed under the regulation of the T7 promoter in Escherichia coli. PAC, with its complex formation mechanism, serves as a unique target protein for demonstration of several key strategies for enhancing recombinant protein production. The current T7 system for pac overexpression was fraught with various technical hurdles. Upon the induction with a conventional inducer of isopropyl-β-d-thiogalactopyranoside (IPTG), the production of PAC was limited by the accumulation of PAC precursors (proPAC) as inclusion bodies and various negative cellular responses such as growth inhibition and cell lysis. The expression performance could be improved by the coexpression of degP encoding a periplasmic protein with protease and chaperone activities. In addition to IPTG, arabinose was shown to be another effective inducer. Interestingly, arabinose not only induced the current T7 promoter system for pac expression but also facilitated the posttranslational processing of proPAC for maturation, resulting in significant enhancement for the production of PAC. Glycerol appeared to have an effect similar to, but not as significant as, arabinose for enhancing the production of PAC. The study highlights the importance of developing suitable genetically engineered strains with culture conditions for enhancing recombinant protein production in E. coli.

Keywords

Inclusion Body Arabinose Recombinant Protein Production Enhance Cell Growth lacUV5 Promoter 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgements

This study was supported in part by the Natural Sciences and Engineering Research Council of Canada and National Science Council of Taiwan.

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Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Yali Xu
    • 1
  • Stefan Rosenkranz
    • 1
  • Chiao-Ling Weng
    • 2
  • Jeno M. Scharer
    • 1
  • Murray Moo-Young
    • 1
  • C. Perry Chou
    • 1
    Email author
  1. 1.Department of Chemical EngineeringUniversity of WaterlooWaterlooCanada
  2. 2.Department of Chemical EngineeringFeng Chia UniversityTaichungRepublic of China

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