Applied Microbiology and Biotechnology

, Volume 103, Issue 8, pp 3393–3406 | Cite as

Soluble expression of IGF1 fused to DsbA in SHuffle™ T7 strain: optimization of expression and purification by Box-Behnken design

  • Nabbi Emamipour
  • Manouchehr Vossoughi
  • Fereidoun Mahboudi
  • Majid GolkarEmail author
  • Pezhman Fard-EsfahaniEmail author
Biotechnological products and process engineering


Production of insulin-like growth factor 1 (IGF1) in Escherichia coli mostly results in the formation of inclusion bodies. In the present study, IGF1 was fused to disulfide bond oxidoreductase A (DsbA) and expressed in SHuffle™ T7 strain, in order to obtain correctly folded protein. Soluble expression and IMAC purification of DsbA-IGF1 were optimized by applying the Box-Behnken design of response surface methodology. The optimization greatly increased concentration of soluble protein from 317 to 2600 mg/L, and IMAC yield from 400 to 1900 mg/L. Results of ANOVA showed induction OD600 and temperature had significant effects on the soluble protein expression while isopropyl-β-d thiogalactoside, in the concentrations tested, displayed no significant effect. Moreover, the three parameters of the binding buffer including, pH, concentration of NaCl, and imidazole displayed significant effects on the IMAC yield. Then, purified DsbA-IGF1 was cleaved by human rhinovirus 3C protease, and authentic IGF1 was obtained in flow through of a subtractive IMAC. Final polishing of the protein by reversed-phase HPLC yielded IGF1 with purity of 96%. The quality attributes of purified IGF1 such as purity, identity, molecular size, molecular weight, secondary structure, and biological activity were assessed and showed to be comparable to the standard IGF1. The final yield of purified IGF1 was estimated to be 120 ± 18 mg from 1 L of the culture. Our results demonstrated a simple and easily scalable strategy for production of large amounts of bioactive IGF1 by rational designing soluble protein expression, and further optimization of expression and purification methods.


Insulin-like growth factor 1 Soluble expression HPLC E. coli Protein purification Box-Behnken design 



The present study was a part of PhD thesis of N. Emamipour, and was funded by Pasteur Institute of Iran (grant number 95/0110/2542).

Compliance with ethical standards

Conflict of Interest

The authors declare that they have no competing interests.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Molecular Parasitology Laboratory, Department of ParasitologyPasteur Institute of IranTehranIran
  2. 2.Department of BiochemistryPasteur Institute of IranTehranIran
  3. 3.Department of Chemical and Petroleum EngineeringSharif University of TechnologyTehranIran
  4. 4.Biotechnology Research CenterPasteur Institute of IranTehranIran

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