An efficient large-scale refolding technique for recovering biologically active recombinant human FGF-21 from inclusion bodies

Abstract

Fibroblast growth factor 21 (FGF-21) is an important regulator in glycolipid metabolism that is a promising drug candidate for treatment of diabetes and obesity. However, the productivity of recombinant hFGF-21 (rhFGF-21) in Escherichia coli (E. coli) is relatively low, which limits its clinical application. To meet the clinical demand and control the production cost, rhFGF-21 proteins were expressed in inclusion bodies (IBs) form in Rosetta (DE3) by high cell density fermentation in 50-L scale. Hollow fiber membrane filtration technology was used to enrich the bacteria, wash, denature and refold the IBs in the current report. The renatured proteins were purified by two-step affinity chromatography. Authenticity of the purified rhFGF-21 was confirmed by the N-and C-terminal sequence, disulfide bond composition and molecular weight analyses. Results showed that the average target protein and recovery of rhFGF-21 expressed in IBs form of three batches were more than those of the soluble form. Both the rhFGF-21 proteins from the two forms showed equal potency in improving the glucose uptake in HepG2 cells and anti-diabetic effect in db/db mice. In this study, an efficient method for preparation of FGF-21 was established. This novel process provides an important technical basis for the large-scale production of rhFGF-21.