Isolation, Purification And Identification Of Recombinant Human Hepcidin

Hepcidin is a low-molecular-weight, highly disulfide bonded peptide relevant to small intestine iron absorption and body iron homeostasis. Like other cysteine-rich antimicrobial peptides, hepcidin also exhibits obvious antibacterial and antifungal activity. Method of isolation and purification of recombinant hepcidin is described, and the bioactivity of the protein was assayed in this paper.The 10.6 kDa His-hepcidin expressed from pET-hpc was exitsted in the form of insoluble inclusion bodies. Then the inclusion bodies was solubilized in 8mol/L urea containing 100mmol/L 2-ME and 50mmol/L Tris-HCl(pH 8.0).Immobilized metal ion affinity chromatography (IMAC) was used to purify His-hepcidin in the condition of denaturation with two step elution. Firstly, 60mmol/L imidazole(pH 8.0) was used to elute the contaminants from the column. Then the elution of His-hepcidin was performed at pH 4.0. The binding capacity of Ni²⁺-IDA Sepharose Fast Flow for His-hepcidin was 30.4mg/ml resin.