Improvement of yield and purity of human fibroblast growth factor-21.
- Author:
Dan YU
;
Xianlong YE
;
Guiping REN
;
Pengfei XU
;
Shujie LI
;
Zeshan NIU
;
Deshan LI
- Publication Type:Journal Article
- MeSH:
Animals;
Bacteria;
metabolism;
Diabetes Mellitus, Experimental;
drug therapy;
Disease Models, Animal;
Fibroblast Growth Factors;
biosynthesis;
Genetic Vectors;
Glucose;
metabolism;
Hep G2 Cells;
Humans;
Hypoglycemic Agents;
isolation & purification;
Inclusion Bodies;
metabolism;
Mice;
Recombinant Fusion Proteins;
biosynthesis;
Small Ubiquitin-Related Modifier Proteins;
biosynthesis
- From:
Chinese Journal of Biotechnology
2014;30(4):658-668
- CountryChina
- Language:Chinese
-
Abstract:
Fibroblast growth factor -21 (FGF-21) is a recently discovered metabolic regulation factor, regulating glucose and lipid metabolism and increasing insulin sensitivity. FGF-21 is expected to be a potential anti-diabetic drug. Expression of FGF-21 as inclusion bodies has advantages for high yield and purity, but the bioactivity of the protein is almost totally lost after denature and renature. That is why FGF-21 is currently expressed in soluble form. As a result, the yield is considerably low. In this study, we used SUMO vector to express SUMO-human FGF-21 (SUMO-hFGF-21) in form of inclusion body. We optimized the culture conditions to increase the yield of the bioactive human fibroblast growth factor-21. We applied the hollow fiber membrane filtration column to enrich the bacteria, wash, denature and renature inclusion bodies. After affinity and gel filtration chromatography, we examined the hypoglycemic activity of FGF-21 by the glucose uptake assay in HepG2 cells. We also detected the blood glucose concentration of type 2 diabetic db/db model mice after short or long-term treatment. The results show that the yield of ihFGF-21 was 4 times higher than that of shFGF-21. The yield was 20 mg/L for ihFGF-21 vs. 6 mg/L for shFGF-21. The purity of ihFGF-21 was above 95%, while shFGF-21 was 90%. Compared with the traditional method of extracting inclusion bodies, the production cycle was about three times shortened by application of hollow fiber membrane filtration column technology, but the bioactivity did not significantly differ. This method provides an efficient and cost-effective strategy to the pilot and industrial production of hFGF-21.
