Preparation and identification of functional recombinant human macrophage migration inhibitory factor

Macrophage migration inhibitory factor(MIF),as a specific cytokine,possesses hormonelike efficacy.In this study, we try to prepare the recombinant human macrophage migration inhibitory factor(hMIF) with bioactivity and promote the research of MIF activity inhibitor.We constructed prokaryotic expression vectors PET11b-MIF and PET32a-MIF and established purifying methods for them The bioactivity of these two recombinant proteins was compared by testing tautomerase activity,stimulating the release of inflammatory cytokine,and increasing the mortality of sepsis mice.We successfully constructed two recombinant hMIF,as well as established and optimized the methods to purify recombinant hMIF.After purified with SP Sepharose FF cation-exchange chromatography and Hitrap 26/60 sephacryl S-300 gel filtration chromatograph,MIF without any label has obvious tautomerase activity,and could stimulate RAW264.7 cells to release NO,TNF-α,IL-6 and other inflammatory factors in a dose-dependent manner,and also can increase mortality of sepsis mice.But the MIF with Trx tag(Trx-MIF) showed no such effects.These results indicated that the papered functional MIF with bioactivity would be useful to explore the function of MIF and promote the research of MIF activity inhibitor.