Optimization of Production Acetyl xylan Esterase in Pichia postoris and Its Synergy with Xylanase in Xylan Hydrolysis

摘要

A new Volvariella volvacea gene encoding an acetyl xylan esterase (designated as axe1) was cloned and expressed in Pichia pastoris. To further increase the production of AXE1 by yeast strains KM71H (Muts), the factors which affected acetyl xylan esterase production in baffled flask were screened out by single factor analysis. The result showed that three prominent factors were methanol concentration, the amount of casein added, and initial pH in the medium. Then, Box Behnken design and Response Surface Methodology was employed for optimizing the fermentation condition. The optimal fermentation conditions were methanol 2.8%, casein 0.63%, initial pH 8.0, initial cell density OD600 30, and temperature 25. Under these conditions, the yield ℃ of AXE1 increased from 209IU/ml to 463IU/ml. The production of AXE1 was further incresed by high cell-density fermentation in a 7.5L fermenter. The activity of enzyme reached 1385 IU/mL after 84 hours’ fermentation in the best condition casein 0.65%, pH 6.5 of induction phase, and methanol concentration 0.35% -0.5%, respectively. The purified AXE1 showed a mild but significant synergistic effect in combination with crude xylanase from Volvariella volvacea. On the degradation of oat-spelt xylan, birchwood xylan, and DSWB. A 1.04, 1.11 and 1.01-fold increases in the amount of reducing sugar were released, compared to the expected amount of individual enzymes alone. This indicates that the acetyl xylan esterase has potential industrial applications in the utilizing renewable hemicelluloses containing acetyl xylan.