Volume 5 Supplement 1

The 4th Recombinant Protein Production Meeting: a comparative view on host physiology

Open Access

Addition of Repressor in inducible promoter system improves soluble expression of recombinant protein in E. coli

  • Kyung-Hwan Jung1
Microbial Cell Factories20065(Suppl 1):P19

DOI: 10.1186/1475-2859-5-S1-P19

Published: 10 October 2006

Background

Insoluble protein aggregate (inclusion body) is frequently accumulated during the heterologous protein expression by the bacterial inducible promoter system. In this study, although many reports have proposed the methodology to circumvent the aggregate formation [13], we tried to control the transcription rate by an addition of the repressor for inducible promoter. The addition of repressor was tried just after the inducer was added, in order to increase the soluble expression level.

Results

To improve the soluble expression level of recombinant interferon-alpha (IFN-alpha) in E. coli, repressor (glucose) was added after induction. In this system, arabinose-inducible promoter (pBAD) controlled the transcription of IFN-alpha gene. The fractionation of soluble and insoluble part of the induced E. coli by B-PERII solution (Pierce) showed that glucose addition after induction resulted in improvement of the soluble expression, otherwise IFN-alpha was expressed mostly in insoluble portion (see Figure 1). Finally, over 60% of the total protein expression was found in the soluble fraction of total cell lysate. Probably this principle might be able to apply to other heterologous protein expression which is prone to a protein aggregate formation in the cytoplasm.
Figure 1

Western blot analysis; (A) Soluble and insoluble expression of recombinant interferon-alpha by arabinose induction (0.05%), (B) Soluble and insoluble expression of recombinant interferon-alpha by arabinose induction (0.05%) and 0.5% glucose addition after induction.

Conclusion

The glucose (repressor) addition improved the soluble expression level in arabinose-inducible promoter system in E. coli.

This principle might be able to apply to a heterologous protein expression which is prone to a protein aggregate formation in the cytoplasm.
Figure 2

The change of soluble and insoluble fraction after arabinose induction. Glucose (repressor) was added after induction (,). In other two cases (,□), glucose was not added. Fraction was obtained from image analysis of Figure 1.

Authors’ Affiliations

(1)
Department of Food and Biotechnology, Chungju National University

References

  1. Baneyx F: vivo folding of recombinant proteins in Escherichia coli. Manual of Industrial Microbiology and Biotechnology. Edited by: Demain AL, et al. 1999, ASM, Google Scholar
  2. Swartz JR: Advances in Escherichia coli production of therapeutic proteins. Curr Opin Biotechnol. 2001, 12: 195-201. 10.1016/S0958-1669(00)00199-3.View ArticleGoogle Scholar
  3. Sorensen HP, Mortensen KK: Soluble expression of recombinant protein in the cytoplasm of Escherichia coli . Microb Cell Fact. 2005, 4: 1-. 10.1186/1475-2859-4-1.View ArticleGoogle Scholar

Copyright

© Jung; licensee BioMed Central Ltd. 2006

This article is published under license to BioMed Central Ltd.

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