特异腐质霉角质酶在大肠杆菌中的表达和发酵优化  

Expression of Humicola insolens Cutinase in Escherichia coli and its Optimization of Fermentation

作　　者：孙益荣 吴敬[1,2,3] 宿玲恰 SUN Yi-rong;WU Jing;SU Ling-qia(State Key Laboratory of Food Science and Technology Jiangnan University, Wuxi, Jiangsu 214122, China;School of Biotechnology and Key Laboratory of Industrial Biotechnology Ministry of Education Jiangnan University, Wuxi , Jiangsu 214122, China;International Cooperation Laboratory of Food Science and Technology of the Ministry of Education, Wuxi , Jiangsu 214122, China)

机构地区：[1]江南大学食品科学与技术国家重点实验室,江苏无锡214122 [2]江南大学生物工程学院工业生物技术教育部重点实验室,江苏无锡214122 [3]江南大学教育部食品安全国际合作联合实验室,江苏无锡214122

出　　处：《食品与机械》2018年第4期1-5,共5页Food and Machinery

基　　金：国家杰出青年基金(编号:31425020)

摘　　要：应用大肠杆菌(Escherichia coli,E.coli)BL21(DE3)表达系统实现特异腐质霉(Humicola insolens,H.insolens)来源角质酶的重组表达。对重组角质酶进行酶学性质研究,发现其最适pH为8.5、最适温度为80℃,并有良好的pH稳定性和温度稳定性。经3L发酵罐中扩大培养并进行发酵条件优化,其最佳发酵条件:发酵前期控制菌体生长pH 7.0、温度37℃,培养时间12~14h;菌体浓度OD600达到50时进入诱导期,调节温度降至30℃,恒速流加0.2g/(L·h)的乳糖溶液进行诱导,总发酵周期为36h,最高酶活2 233U/mL,是摇瓶水平酶活的13倍。Cutinase is a kind of multifunctional hydrolase,which has important application value in the food processing and textile refining industry.The recombinant expression system of Escherichia coli（E.coli）BL21（DE3）was used to achieve the recombinant expression of Humicola insolens cutinase in E.coli, and the activity of extracellular supernatant in shaking flask level reached 170 U/mL.The recombinant cutinase was characterized.The optimum pH was 8.5,the optimum temperature was 80℃,with good pH stability and temperature stability.In order to further improve the possibility of the industrial application of H.insolens cutinase,the recombinant E.coli was cultured in 3 Lfermentor and the cultivation condition is optimized.The optimum fermentation conditions：maintain pH 7.0 and 37 ℃in early culture phases about 12~14 h;when the cell concentration OD600 up to 50,fermentation entered induced phases,adjust the temperature to 30 ℃,0.2 g/（L·h）lactose solution were added with constant flow rate.The total fermentation period was about 36 h.The highest activity of enzyme reached to 2 233 U/mL,which is 13 times about the enzyme activity in shake flask level.

关 键 词：特异腐质霉 角质酶 重组大肠杆菌 发酵优化 

分 类 号：TQ925[轻工技术与工程—发酵工程]