Optimization of conditions for expression of recombinant interferon-γ in E.coli

Interferon gamma (IFN-gamma) is an important immunoregulatory cytokine that has a central role against viral and bacterial infections. In this study, the cDNA encoding 141 amino acids of mature IFN-gamma from mice splenocytes was cloned in a prokaryotic expression vector pQE 30. Optimization of expression conditions resulted in high IFN-gamma protein. Western blot showed that recombinant IFN-gamma was specifically recognized by its counterpart anti-mouse IFN-gamma antibodies. In vitro dose-dependent studies, with A549 and HeLa cell lines, showed that cloned IFN-gamma was safe and had no effect on cell proliferation. The protein prediction and analysis using SOPMA program, revealed that IFN-gamma had 80 alpha-helices, 8 beta-turns jointed by 9 extended strands and 44 random coils. A total of four major clusters were observed with murine IFN-gamma sharing 39 % homology with human IFN-gamma. Pair-wise alignment studies with human revealed 26 % identity and 43.3 % similarity. The recovery of bioactive proteins from inclusion bodies (IBs) is a complex process and various protocols have been developed. We report here a simple, robust and inexpensive purification approach for obtaining recombinant IFN-gamma protein expressed as IBs in E.coli.