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Optimization of an induction strategy for improving interferon‐α2b production in the periplasm of Escherichia coli using response surface methodology
Author(s) -
Azaman Siti Nor Ani,
Ramakrishnan Nagasundara Ramanan,
Tan Joo Shun,
Rahim Raha Abdul,
Abdullah Mohd Puad,
Ariff Arbakariya B.
Publication year - 2010
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1042/ba20100104
Subject(s) - periplasmic space , escherichia coli , recombinant dna , lac operon , inclusion bodies , chemistry , biology , biochemistry , microbiology and biotechnology , gene
Induction strategies for the periplasmic production of recombinant human IFN‐α2b (interferon‐α2b) by recombinant Escherichia coli Rosetta‐gami 2(DE3) were optimized in shake‐flask cultures using response surface methodology based on the central composite design. The factors included in the present study were induction point, which related to the attenuance of the cell culture, IPTG (isopropyl β‐ D ‐thiogalactoside) concentration and induction temperature. Second‐order polynomial models were used to correlate the abovementioned factors to soluble periplasmic IFN‐α2b formation and percentage of soluble IFN‐α2b translocated to the periplasmic space of E. coli . The models were found to be significant and subsequently validated. The proposed induction strategies consisted of induction at an attenuance of 4 (measured as D 600 ), IPTG concentration of 0.05 mM and temperature of 25 °C. The optimized induction strategy reduced inclusion‐body formation as evidenced by electron microscopy and yielded 323.8 ng/ml of IFN‐α2b in the periplasmic space with translocation of 74% of the total soluble product. In comparison with the non‐optimized condition, soluble periplasmic production and the percentage of soluble IFN‐α2b translocated to the periplasmic space obtained in optimized induction strategies were increased by approx. 20‐fold and 1.4‐fold respectively.