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Cloning, Expression, Purification and Evaluation of the Biological Properties of the Recombinant Human Growth Hormone (hGH) in Escherichia coli

  • A. Doozandeh-Juibari
  • S. GhovvatiEmail author
  • H. R. Vaziri
  • M. M. Sohani
  • Z. Pezeshkian
Article
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Abstract

The 22 kDa of human growth hormone (hGH) is naturally produced and secreted by somatotrophic cells in the anterior part of the pituitary gland. The aim of this study was to clone, express, purify of hGH as fusion to the pelB leader in pET22b (+) plasmid and evaluate it’s biological properties. The hGH polypeptide codon was optimized and subcloned. The recombinant hGH protein was purified by affinity chromatographic system against His-tag and the presence and accuracy of the purified products were evaluated by protein electrophoresis and Western blot. The biological activity of recombinant hGH protein was measured using ELISA assay. The results of sequencing of hGH gene confirmed the presence and proper placement of hGH gene and it’s subcloning in the plasmid pET22b (+). The results of the protein electrophoresis and Western blot assays demonstrated that the expression accuracy of 22 kDa recombinant hGH. The results of Bradford spectroscopy assay showed that the recombinant hGH protein concentration was 1 g/l. The results of classical sandwich ELISA assay, in contrast to the specific antibodies, confirmed the bio-activity of the recombinant hGH protein in its targeting. Consequently, the results of this study showed that pelB leader has the ability to more accurately direct hGH to periplasmic space in Escherichia coli, and the conditions of oxidizing periplasmic space give rise to the correct folding of the protein in this space. Furthermore, the results of current study proved that using bioinformatics tools and combining them with laboratory data, could improve the recombinant hGH expression in E. coli, in addition to preserving bio-activity.

Keywords

Human growth hormone (hGH) Recombinant protein Cloning and expression ELISA Affinity chromatography Western blot 

Notes

Funding

This study was no funded.

Compliance with Ethical Standards

Conflict of interest

The authors declare that we have no conflict of interest.

Ethical Approval

No human or animals subjects were involved in this study.

Informed Consent

This study contain no individual participant data.

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • A. Doozandeh-Juibari
    • 1
  • S. Ghovvati
    • 2
    Email author
  • H. R. Vaziri
    • 1
  • M. M. Sohani
    • 2
  • Z. Pezeshkian
    • 3
  1. 1.Department of Molecular Cell Biology, College of Applied ScienceUniversity of GuilanRashtIran
  2. 2.Department of Biotechnology, Faculty of AgricultureUniversity of GuilanRashtIran
  3. 3.Department of Animal Sciences, Faculty of AgricultureUniversity of GuilanRashtIran

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