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

Matsui, Toru; Togari, Takashi; Misawa, Satoru; Namihira, Tomoyuki; Shinzato, Naoya; Matsuda, Hitoshi; Sato, Seigo

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.

ESCHERICHIA coli; AMINO acids; PLASMIDS; CELL culture; GENE expression

Applied Biochemistry & Biotechnology, 2010, Vol 162, Issue 3, p823

0273-2289

Academic Journal

10.1007/s12010-010-8913-4