Extracellular expression and antiviral activity of a bovine interferon-alpha through codon optimization in Pichia...

Interferons (IFNs) are the primary line of defense against infectious agents. In particular, IFN-α is an important antiviral cytokine and has a wide range of immune-modulating functions. Porcine and human IFN-α have been successfully prepared and play important roles in the prevention and therapy of viral diseases. To date, there has been limited applied research on bovine IFN-α. To achieve high-level expression of recombinant bovine IFN-α (bIFN-α) in Pichia pastoris for large-scale application, the bIFN-α gene was optimized and synthesized on the basis of codon bias of P. pastoris. Optimized bIFN-α (opti-bIFN-α) was successfully expressed in P. pastoris and directly secreted into the culture supernatant. The amount of extracellular soluble opti-bIFN-α was observed to be 200μg/mL in a shake flask. Expression efficiency of opti-bIFN-α was found to be about three times that of wild-type bIFN-α when the expression yield was compared at the same copies of the targeted gene. In addition, both the original cultural supernatant and purified opti-bIFN-α showed strong antiviral activity in MDBK cells (2×10(6)AU/mL and 1×10(7)AU/mg, respectively) and IBRS-2 cells (3×10(5)AU/mL and 1.5×10(6)AU/mg, respectively) against a recombinant vesicular stomatitis virus expressing the green fluorescence protein. In this study, we demonstrated high-level extracellular expression of opti-bIFN-α by P. pastoris. To the best of our knowledge, the opti-bIFN-α yield observed in this study is the highest to be reported to date. Our results demonstrated that the extracellular opti-bIFN-α with strong antiviral activity could be easily prepared and purified at a low cost and that it may be a potential biological therapeutic drug against bovine viral infections.