Enhanced porcine interferon-α production by Pichia pastoris by methanol/sorbitol co-feeding and energy metabolism shift

Abstract:

Porcine interferon-α (pIFN-α) fermentative production by recombinant Pichia pastoris was carried out in a 10-L bioreactor to study its metabolism changes and effects on fermentation under different inducing strategies, by analyzing the change patterns of the corresponding metabolism and energy regeneration. The results show that the specific activities of alcohol oxidase (AOX), formaldehyde dehydrogenase (FLD) and formate dehydrogenase (FDH) largely increased when reducing temperature from 30 °C to 20 °C under pure methanol induction, leading significant enhancements in methanol metabolism, formaldehyde dissimilatory energy metabolism and pIFN-α antiviral activity. The highest pIFN-α antiviral activity reached 1.4×106 IU/mL, which was about 10-folds of that obtained under 30 °C induction. Using methanol/sorbitol co-feeding strategy at 30 °C, the major energy metabolism energizing pIFN-α synthesis shifted from formaldehyde dissimilatory energy metabolism pathway to TCA cycle, formaldehyde dissimilatory pathway was weakened and accumulation of toxic intermediate metabolite-formaldehyde was relieved, and methanol flux distribution towards to pIFN-α synthesis was enhanced. Under this condition, the highest pIFN-α antiviral activity reached 1.8×107 IU/mL which was about 100-folds of that obtained under pure methanol induction at 30 °C. More important, enhanced pIFN-α production with methanol/sorbitol co-feeding strategy could be implemented under mild conditions, which greatly reduced the fermentation costs and improved the entire fermentation performance.