Influence of methanol concentration on purification recovery of consensus interferon-a produced by Pichia pastoris / 生物工程学报

ABSTRACT

We observed the influence of methanol concentration on purification recovery of recombinant human consensus interferon-a (cIFN) produced by Pichia pastoris. Fermentations controlled at 0.75% (W/V) methanol showed a 24% increase in clFN expression compared to using 0.25% methanol. However, the purification recovery rate of cIFN at 0.25% methanol was 3.75-fold higher than that at 0.75% methanol. To seek the reason, we analyzed the stability of clFN by SDS-PAGE and Native-PAGE as well as Western blotting. The electrophoresis results revealed that cIFN formed a lot of aggregates in media when the induction was controlled at 0.75% methanol, and two different aggregate forms were found disulfide bond covalent aggregates and non-covalent aggregates. However, these aggregates almost disappeared when the methanol concentration was controlled at 0.25%, at the same time, cIFN bioactivity of supernatant increased almost 4.48-fold. Finally, 0.73g monomer cIFN was obtained after purification from 1 liter supernatant at 0.25% methanol induction, showed a 2.84-fold increase compare to the induction at 0.75% methanol.