Highly efficient soluble expression and purification of recombinant human basic fibroblast growth factor (hbFGF) by fusion with a new collagen-like protein (Scl2) in Escherichia coli

• Published in: Preparative biochemistry & biotechnology Vol. 50; no. 6; pp. 598 - 606
• Main Authors: Rahman, Inamur, Fang, Lina, Wei, Zhang, Zheng, Xiaodong, Jiazhang, Lian, Huang, Lei, Xu, Zhinan
• Format: Journal Article
• Language: English
• Published: England Taylor & Francis 02.07.2020; Taylor & Francis Ltd
• Subjects: Amino Acid Sequence; Animals; Bacterial collagen-like protein; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; BALB 3T3 Cells; bioactive properties; Biological activity; biotechnology; carbon; Carbon sources; Cell density; Cell Proliferation - drug effects; Chemical compounds; Chromatography, Ion Exchange; Cloning, Molecular - methods; Collagen; Collagen - genetics; Collagen - metabolism; cost effectiveness; culture flasks; E coli; enteropeptidase; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; fermenters; Fibroblast growth factor 2; Fibroblast Growth Factor 2 - genetics; Fibroblast Growth Factor 2 - metabolism; Fibroblasts; Fibroblasts - drug effects; Fibroblasts - metabolism; Freeze drying; fusion expression; Glycerol; Growth factors; Humans; Ion exchange; Ion-exchange chromatography; Mice; Optimization; pH change-dependent acid precipitation; Pharmacology; Production methods; Proteins; Purification; purification methods; Recombinant Fusion Proteins - isolation & purification; Recombinant Fusion Proteins - metabolism; Recombinant Fusion Proteins - pharmacology; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Streptococcus pyogenes; Streptococcus pyogenes - metabolism; therapeutics; Wound healing; fibroblast growth factor-2; Bacterial collagen-like protein; fusion expression; Escherichia coli; pH change-dependent acid precipitation
• ISSN: 1082-6068; 1532-2297; 1532-2297
• DOI: 10.1080/10826068.2020.1721533

Human basic fibroblast growth factor (hbFGF) is involved in a wide range of biological activities that affect the growth, differentiation, and migration. Due to its wound healing effects and therapy, hbFGF has the potential as therapeutic agent. Therefore, large-scale production of biologically active recombinant hbFGF with low cost is highly desirable. However, the complex structure of hbFGF hinders its high-level expression as the soluble and functional form. In the present study, an efficient, cost-effective, and scalable method for producing recombinant hbFGF was developed. The modified collagen-like protein (Scl2-M) from Streptococcus pyogenes was used as the fusion tag for producing recombinant hbFGF for the first time. After optimization, the expression level of Scl2-M-hbFGF reached approximately 0.85 g/L in the shake flask and 7.7 g/L in a high cell-density fermenter using glycerol as a carbon source. Then, the recombinant Scl2-M-hbFGF was readily purified using one-step acid precipitation and the purified Scl2-M-hbFGF was digested with enterokinase. The digested mixture was further subject to ion-exchange chromatography, and the final high-purity (96%) hbFGF product was prepared by freeze-drying. The recovery rate of the whole purification process attained 55.0%. In addition, the biological activity of recombinant hbFGF was confirmed by using L929 and BALB/c3T3 fibroblasts. Overall, this method has the potential for large scale production of recombinant hbFGF.