Improved Production of Aspergillus usamii endo-β-1,4-Xylanase in Pichia pastoris via Combined Strategies

Abstract EN

A series of strategies were applied to improve expression level of recombinant endo-β-1,4-xylanase from Aspergillus usamii (A.

usamii) in Pichia pastoris (P.

pastoris).

Firstly, the endo-β-1,4-xylanase (xynB) gene from A.

usamii was optimized for P.

pastoris and expressed in P.

pastoris.

The maximum xylanase activity of optimized (xynB-opt) gene was 33500 U/mL after methanol induction for 144 h in 50 L bioreactor, which was 59% higher than that by wild-type (xynB) gene.

To further increase the expression of xynB-opt, the Vitreoscilla hemoglobin (VHb) gene was transformed to the recombinant strain containing xynB-opt.

The results showed that recombinant strain harboring the xynB-opt and VHb (named X33/xynB-opt-VHb) displayed higher biomass, cell viability, and xylanase activity.

The maximum xylanase activity of X33/xynB-opt-VHb in 50 L bioreactor was 45225 U/mL, which was 35% and 115% higher than that by optimized (xynB-opt) gene and wild-type (xynB) gene.

Finally, the induction temperature of X33/xynB-opt-VHb was optimized in 50 L bioreactor.

The maximum xylanase activity of X33/xynB-opt-VHb reached 58792 U/mL when the induction temperature was 22°C.

The results presented here will greatly contribute to improving the production of recombinant proteins in P.

pastoris.