重组毕赤酵母发酵生产α-葡聚糖酶的放大研究

Scale-up Fermentation of Recombinant Dextranase Expressed in Pichia Pastoris

研究了毕赤酵母高密度发酵组成型表达α-葡聚糖酶,从6.8 L发酵罐逐级放大至5000 L发酵罐的放大工艺。采用恒定p H和控制甘油残留浓度和溶解氧相结合的调控策略,进行了多批次的发酵试验。结果表明:三磷酸甘油醛脱氢酶启动子驱动的α-葡聚糖酶的生成具有生长偶联特征,集中在对数生长期和稳定期。6.8 L发酵罐中上清酶活达1253 U/m L,中试放大至50 L发酵罐中产酶最高达1300 U/m L,500 L发酵罐中酶活为740 U/m L。5000 L规模发酵罐中产酶达到589 U/m L。建立了以甘油作为碳源,发酵调控简单的工艺,避免了使用甲醇带来的安全问题,适合放大生产。

The high density fermentation of dextranase in the constitutive Pichia pastoris expression system scaled up from 6.8 to 5000 L in series was studied. By controlling residual concentration of glycerol and dissolved oxygen combined with p H-stat, multi batches of fermentation were performed. Results showed that the dextranase synthesis under GAP promoter is growth-associated and mainly in exponential phase and stationary phase. Activity of dextranase in 6.8 L, 50 L, 500 L and 5000 L fermenter reached 1253, 1300, 740 and 589 U/m L, respectively. A simple and convenient fermentation method using glycerol as carbon source has been developed, which obviates problems related with methanol safety concerns and is suitable for large scale production.