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Applied Biochemistry and Biotechnology

, Volume 162, Issue 3, pp 823–829 | Cite as

Optimized Culture Conditions for the Efficient Production of Porcine Adenylate Kinase in Recombinant Escherichia coli

  • Toru MatsuiEmail author
  • Takashi Togari
  • Satoru Misawa
  • Tomoyuki Namihira
  • Naoya Shinzato
  • Hitoshi Matsuda
  • Seigo Sato
Article

Abstract

Temperature shift cultivations with amino acid supplementation were optimized to produce porcine adenylate kinase (ADK) in recombinant Escherichia coli harboring a pUC-based recombinant plasmid under the control of the trp promoter. With regard to temperature control, the culture condition was initially maintained at 35 °C for cellular growth, but ADK expression was suppressed until the late logarithmic growth phase; subsequently, a temperature shift was applied (from 35 °C to 42 °C), which resulted in maximal ADK production. In addition, supplementation of amino acids, especially valine and leucine, during the temperature shift stimulated ADK expression from 3.5% to 9.2% and 8.6% of the total protein, respectively. After optimization, 1 g ADK per liter was produced within 16 h of cultivation with a dry cell weight of 21.8 g/l. In this system, there was no loss of the recombinant plasmid during cultivation without selective pressure.

Keywords

High cell density cultivation Escherichia coli Porcine adenylate kinase Temperature shift cultivation 

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Toru Matsui
    • 1
    Email author
  • Takashi Togari
    • 2
  • Satoru Misawa
    • 3
  • Tomoyuki Namihira
    • 1
  • Naoya Shinzato
    • 1
  • Hitoshi Matsuda
    • 4
  • Seigo Sato
    • 2
  1. 1.Center of Molecular BiosciencesUniversity of the RyukyusOkinawaJapan
  2. 2.Graduate School of Life and Environmental SciencesUniversity of TsukubaTsukuba-shiJapan
  3. 3.Mitsubishi Chemical Co.YokohamaJapan
  4. 4.Bio-Research CentreJapan Energy CoSaitamaJapan

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