为了考察重组地衣芽孢杆菌的异源表达特性,进一步提高麦芽四糖酶的发酵水平,以适用于工业化生产,研究构建了木糖诱导型的重组地衣芽孢表达系统,表达了来源于嗜糖假单胞菌的麦芽四糖酶基因,并进行了初步发酵优化。结果显示,构建的表达系统经过核酸电泳以及十二烷基硫酸钠-聚丙烯酰氨凝胶电泳(sodium dodecyl sulfate-polyacrylamide gel electrophoresis,SDS-PAGE)鉴定均正确,该系统在发酵过程中经过木糖诱导,重组酶能够成功地分泌至发酵液中,然后通过对诱导剂添加时间、诱导温度和发酵时间优化后,摇瓶水平的最大酶活力可达(168.2±9.41) U/mL。最佳的发酵条件为,在24 h添加诱导剂,诱导温度30 ℃,发酵至稳定期结束停止发酵。该研究为重组地衣芽孢杆菌的异源表达研究以及麦芽四糖酶的工业化应用提供了参考。
To investigate the heterologous expression in Bacillus licheniformis and improve the fermentation level of maltotetraose-forming amylase, a xylose-inducible expression system was constructed in B. licheniformis CICIM B1391 to express the maltotetraose-forming amylase from Pseudomonas saccharophila, and a preliminary fermentation optimization was conducted. It was observed that maltotetraose-forming amylase was successfully secreted into the fermentation broth induced by xylose. After optimized induction time, temperature and fermentation time, the maximum enzyme activity reached to (168.2±9.41) U/mL in shake flask fermentation. The optimal fermentation conditions were adding the inducer at 24 h, the induction temperature of 30 ℃, and ending fermentation at the end of stable period.
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