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Author Notes:

*E-mail: jgalipe@emory.edu

Conceived and designed the experiments: AP JD JG. Performed the experiments: AP JD. Analyzed the data: AP JD JG. Contributed reagents/materials/analysis tools: AP JD JG. Wrote the paper: AP JG.

The authors have declared that no competing interests exist.

Subjects:

Research Funding:

This work was supported by National Institute of Health (5R01AI093881). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Keywords:

  • Science & Technology
  • Multidisciplinary Sciences
  • Science & Technology - Other Topics
  • REGULATORY B-CELLS
  • COLONY-STIMULATING FACTOR
  • ESCHERICHIA-COLI
  • THERAPEUTIC PROTEINS
  • RECOMBINANT PROTEINS
  • CANCER-IMMUNOTHERAPY
  • FACTOR-XA
  • IN-VIVO
  • GM-CSF
  • FUSOKINE

Maltose-Binding Protein Fusion Allows for High Level Bacterial Expression and Purification of Bioactive Mammalian Cytokine Derivatives

Tools:

Journal Title:

PLoS ONE

Volume:

Volume 9, Number 9

Publisher:

, Pages e106724-e106724

Type of Work:

Article | Final Publisher PDF

Abstract:

Fusokines are chimeric proteins generated by the physical coupling of cytokines in a single polypeptide, resulting in proteins with highly pleiotropic activity and the potential to treat cancer and autoimmune ailments. For instance, the fusokine GIFT15 (GM-CSF and Interleukin 15 Fusion Transgene) has been shown to be a powerful immunosuppressive protein able to convert naì ve B cells into IL-10-producing B cells. To date, the mammalian cell systems used for the expression of GIFT15 allow for secretion of the protein in the culturing media, an inefficient system for producing GMPcompliant fusokines. In this study we report the bacterial expression of bioactive recombinant GIFT15 (rGIFT15). Indeed, there is a constant demand to improve the expression systems for therapeutic proteins. Expression of a maltose-binding protein (MBP) fusion protein efficiently allowed the accumulation of soluble protein in the intracellular milieu. Optimizing the bacterial culture significantly increased the yield of recombinant protein. The biological activity of rGIFT15 was comparable to that of fusokine derived from a mammalian source. This approach led to the production of soluble, endotoxin-free functional protein, averaging 5 mg of rGIFT15 per liter of culture. This process is amenable to scale up for the development of Food and Drug Administration (FDA)-compliant immune-modulatory rGIFT15.

Copyright information:

© 2014 Pennati et al.

This is an Open Access article distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits making multiple copies, distribution of derivative works, distribution, public display, and publicly performance, provided the original work is properly cited. This license requires copyright and license notices be kept intact, credit be given to copyright holder and/or author.

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