Cloning and expression of recombinant human superoxide dismutase 1 (hSOD1) in Bacillus subtilis 1012

Abstract As an important member of the antioxidant defense system in vivo, human Cu, Zn superoxide dismutase (hSOD1) has become a potential therapeutic agent against host diseases, and new strategies were developed to achieve high-yield SOD1 by using efficient production systems, like E. coli, yeast, and other eukaryotic expression systems. However, the above systems have their own limitations, such as the formation of inclusion bodies, endotoxins production, and low secretion efficiency. In this study, the hSOD1 coding sequence gene was amplified from human HepG2 cells, cloned into a pHT43-His expression vector, and transformed into Bacillus subtilis system. After the optimization with different media, temperatures, and inducer (IPTG) concentrations, the recombinant hSOD1 was successfully expressed in Bacillus subtilis 1012. The results show that hSOD1 was produced as a soluble form in Super rich medium with 0.2 mM of IPTG at 37°C after the induction for 24 h. Besides, 20 g/L of lactose also displayed the same inductive effect on hSOD1 expression as that by 0.2 mM of IPTG. Finally, hSOD1 was efficiently purified by nickel affinity chromatography, and the specific activity of hSOD1 was determined to be 1625 U/mg in the presence of 800 µM of Cu2+ and 20 µM of Zn2+. Collectively, Bacillus subtilis 1012 can not only overcome the shortcomings of E. coli and yeast expression systems, but also be directly used for disease treatment as a healthy probiotic. These advantages provided a possibility for the application of oral engineering bacteria 1012-hSOD1 to the clinical treatment in the future.