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  • Title: Expression and one step intein-mediated purification of biologically active human G-CSF in Escherichia coli.
    Author: Sima S, Shafiee F, Jahanian-Najafabadi A.
    Journal: Mol Biol Rep; 2020 Apr; 47(4):2861-2869. PubMed ID: 32227252.
    Abstract:
    Recombinant form of granulocyte colony stimulating factor (G-CSF) was first approved by FDA in 1998 for chemotherapy induced neutropenia. However, despite production of its biosimilars, less expensive production of G-CSF could reduce the overall therapeutic cost. The aim of this study was to evaluate the possibility of producing biologically active recombinant G-CSF via a single step purification procedure mediated by a self-cleavable intein. G-CSF was expressed by E. coli BL21 (DE3) through IPTG induction, followed by its purification using pH optimization on a chitin column. Western blotting, ELISA, size exclusion chromatography, circular diachorism, peptide mapping, and in vitro assays were performed to compare the structural similarity and biological activity of the purified G-CSF with Neupogen™. Protein purification was confirmed by revealing a band of approximately 18.8 kDa on SDS-PAGE. Bioactivity and physicochemical assays based on the US pharmacopeia showed almost identical or acceptable ranges of similarities between recombinant G-CSF and Neopogen™. this study, biologically active soluble recombinant G-CSF was successfully produced with high purity without using chaotropic solvents through a one-step procedure. This shorter and more efficient purification procedure can reduce the cost and time of G-CSF production which makes its industrial production more cost-effective and might be also applicable for production of other biopharmaceuticals.
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