Purification and Refolding Escherichia coli Produced Enterokinase Inclusion Bodies Immobilized by Nickel Chelating Chromatography

A genetic strain E.coli BL21(DE3),which over-expression enterokinase gene,was constructed.When cell density(OD 600) reached at 0.5-0.6 in LB medium which contained 30 mg/L kanamycin,IPTG was feed to the fermentation broth to 0.4 mmol/L and then temperature changed to 26 oC.After induction for 4 h,the expressed fusion protein concentration was 40% of the total bacteria proteins and in the form of inclusion bodies.The inclusion bodies were further purified under denaturing conditions with affinity of 6 histidine residues at its N-terminal to metal chelate chromographic matrixes.The results showed that the purified recombinant protein under denaturing conditions was refolded in Ni-NTA column by using refolding buffer.The enterokinase activity and purity was 712 U and 96%,respectively.