Enhanced Secretion of Human Interferon Gamma from Bacillus subtilis WB800N

Human interferon gamma (hIFNG) is a 166 amino acid long cytokine having a major role in viral and bacterial diseases like AIDS, Hepatitis, and TB etc. It has been approved and being prescribed as a drug for treatment of chronic granulomatous disease (CGD) and severe malignant osteopetrosis by U.S. FDA. The complete treatment cost is about 37,152 US$. Bacillus subtilis is an emerging GRAS microbial cell factory for extracellular production of recombinant proteins. For toxin free and extracellular IFNG production with low cost downstream processing, we cloned gene encoding human IFNG in expression and extracellular secretion vector, pHT43 and transformed in to Bacillus subtilis WB800N. Sequence and ORF of the cloned gene was confirmed by sequencing and expression was confirmed by antibody based ELISA. The expression of recombinant IFNG under un-optimized conditions was performed in defined minimal and LB medium by inducing the gene with 1mM IPTG at 0h after inoculation and at an OD@600 nm of 0.8. Highest extracellular expression of recombinant IFNG was found to be 600 μgm/l and 300 μgm/l in defined minimal and LB medium, respectively.  The optimal levels of pH, temperature and seed level were determined for enhanced expression of recombinant IFNG.