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

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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
figure1

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
figure2

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.

References

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    Baneyx F: vivo folding of recombinant proteins in Escherichia coli. Manual of Industrial Microbiology and Biotechnology. Edited by: Demain AL, et al. 1999, ASM,

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    Swartz JR: Advances in Escherichia coli production of therapeutic proteins. Curr Opin Biotechnol. 2001, 12: 195-201. 10.1016/S0958-1669(00)00199-3.

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    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.

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Author information

Correspondence to Kyung-Hwan Jung.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution 2.0 International License (https://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Jung, K. Addition of Repressor in inducible promoter system improves soluble expression of recombinant protein in E. coli. Microb Cell Fact 5, P19 (2006) doi:10.1186/1475-2859-5-S1-P19

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Keywords

  • Inclusion Body
  • Protein Aggregate
  • Transcription Rate
  • Inducible Promoter
  • Insoluble Protein