Improvement of the Method of the Expression of rhIL-8 in Escherichia Coli and the Purification

Objective To improve the method of the expression of recombinant human interleukin-8(rhIL-8) in Escherichia Coli and It's purification to obtain high-purified rhIL-8.Method E.Coli was transfected with the constructed rhIL-8 plasmid and the engineering bacterium was induced to express the rhIL-8 by 3-indoleacryl acid.We establish the optimized method of the rhIL-8's expression based on the different concentration of the inducer,temperature,pH and time.Then bacterium was collected by centrifugation and lysed by sonication.Recombinant human IL-8 in the part of inclusion body was recovered by denaturation and renaturation,pooled it with the part of cytoplasm.The pooled fraction was further purified by three steps of chromatography,heparin affinity,ion exchange and gel filtration.We establish the optimized method of purification based on the different condition of chromatography.The purified samples were assessed by SDS PAGE,the quantity of IL-8 was determined by ELISA.Result There were 100 mg/ml rhIL-8 totally when expressing by E.Coli in the condition of 30℃,pH 7.0,50 mmol/L 3-indolecrylic acid,28 hours.The secreting protein,which formed in inclusion body and cytoplasm,was purified to 95% by three steps of chromatography.The IL-8 has good biological activity approved by the test of neutrophil chemotactic.Conclusion High-efficient method of the expression of rhIL-8 in Escherichia Coli and purification method have been established,and the purified IL-8 has good biological activity.