Soluble Expression and Purification of Recombinant Humanized Type I Collagen in Escherichia coli
Article
Figures
Metrics
Preview PDF
Reference
Related
Cited by
Materials
Abstract:
Collagen is currently widely used in the food, medical, and beauty industries. However, isolating recombinant collagens poses considerable challenges because of factors such as inclusion body expression and low purity, limiting its large-scale production and application. Based on humanized type I collagen (hCOL1A1), a recombinant expression vector pET21a(+)-SUMO-hCOL1A1 was constructed by fusing a small ubiquitin-like modifier protein (SUMO) and 6×His tag at the N-terminus of hCOL1A1, which was then transformed into Escherichia coli BL21 (DE3). Soluble expression and purification of recombinant hCOL1A1 in E. coli were achieved through IPTG induction, optimization of the culture conditions, and protein purification of the engineered strain produced by BL21/pET21a(+)-SUMO-hCOL1A1. The supernatant of the engineered strain was purified through Ni-NTA affinity chromatography, TEV enzyme digestion, and ionexchange chromatography, ultimately yielding hCOL1A1 protein with a purity of over 95%. The optimal culture conditions were determined to be an induction temperature of 25 ℃ and a concentration of 0.4 mmol/L IPTG. Further scale-up in 5 L fed-batch fermentation resulted in a final protein yield of 1.43 g/L. Soluble expression of the hCOL1A1 protein, 5 L bioreactor production, and purification were achieved, serving as a valuable reference for the large-scale biosynthesis and purification of recombinant collagen.
