Expression Of Recombinant Porcine Interferon-α In Pichia Pastoris And Its Inhibitional Effect On Virus

[Objective] Pichiapastoris expression system was applied to efficiently express the recombinant porcine interferon-a (rPoIFN-a). The method for purifying rPoIFN-a was established to purify rPoIFN-a from supernatant. The inhibition of rPoIFN-a on Porcine reproductive and respiratory syndrome virus(PRRSV) and Pseudorabies virus(PRV) replication in vitro was tested.[Methods] By the fermentation reactor and pichiapastoris expression system, the expression of rPoIFN-a was induced with methanol in BMMY culture medium. The rPoIFN-a was purified by ammonium sulfate precipitation, Q Sepharose Fast Flow chromatography and gel filtration in turn. The molecular weight and concentration of rPoIFN-a in the expression supernatant and purified one were analysed by Quantity one software. The special activity of rPoIFN-a in the expression supernatant and purified one was determined with Marc-145/VSV system. The antiviral effect of rPoIFN-a on PRRSV and PRV was studied in the present experiment.[Results] The concentration of secreted rPoIFN-a in supernatant reached0.52μg/μL after72hours induction, which accounted for about57.50%of the total amount of protein and its activity was1.00x106u/μL. The purified rPoIFN-a solution contained7.02μg/μL rPoIFN-a with the purity of98.1%and the activity of1.00xl09u/μL. The blocking activity of the purified rPoIFN-a on PRRSV infection prior to treatment with the virus was approximately1.42xl06u/μg. The blocking activity of the purified rPoIFN-a on PRRSV infection prior to treatment with cells was approximately1.42×104u/μg. The replication inhibition effects of the rPoIFN-a on PRRSV was approximately1.42x104u/μg. The blocking activity of the purified rPoIFN-a on PRV infection prior to treatment with the virus or cells was the same, approximately1.42x105u/μg. The replication inhibition effects of the rPoIFN-a on PRV was approximately1.42x103u/μg.[Conclusions] The fermentation test show that Pichia pastoris expression system can largely express rPoIFN-a. The purification method of rPoIFN-a was confirmed, which can highly purify rPoIFN-a. The rPoIFN-a has a high inhibitation effects on PRRSV and PRV infection in vitro.