纤维素酶在酿酒酵母中的表面展示  

Surface display of cellulase on Saccharomyces cerevisiae cells

作　　者：徐小清 刘定康 夏武成 刘子璐 江正兵 XU Xiaoqing;LIU Dingkang;XIA Wucheng;LIU Zilu;JIANG Zhengbing(Hubei Key Laboratory of Industrial Biotechnology,College of Life Sciences,Hubei University,Wuhan 430062,China)

机构地区：[1]湖北大学生命科学学院工业生物技术湖北省重点实验室,湖北武汉430062

出　　处：《湖北大学学报（自然科学版）》2019年第3期228-233,240,共7页Journal of Hubei University：Natural Science

基　　金：湖北省自然科学基金(2018CFA019);湖北省技术创新专项(2018ABA098;2017ABA139);武汉市应用基础研究项目(2017020201010229)资助

摘　　要：利用木质纤维素进行生物法生产燃料乙醇在解决世界能源危机上有十分广阔的前景,但该产业受到预处理的限制,因此以酿酒酵母表面展示技术为依托,构建纤维素酶全细胞酶,既能水解木质纤维素又能利用水解木质纤维素得到的糖类发酵生产乙醇.将编码絮凝素锚定蛋白基因flo1与纤维酶基因cbh2串联整合在同一个开放阅读框中并构建p HBM368-PGK-flo1-cbh2重组表达载体.线性化重组表达载体后再导入尿嘧啶缺陷型酿酒酵母INVSc细胞,经由功能筛选及流式细胞法验证纤维素酶成功地展示表达在酿酒细胞表面.通过DNS法测得该全细胞酶在50℃反应1 h酶活为66. 75 U/mL.成功构建了以酿酒酵母为宿主菌的全细胞酶,为后期全细胞酶协同降解秸秆等系列研究奠定基础.Using lignocellulose to produce fuel ethanol by biological method to solve the world energy crisis has very broad prospects,but it is limited by pretreatment process. Basing on the surface display technology of S. cerevisiae,the whole-cell cellulase was constructed,which could carry out hydrolysis of lignocellulose and ethanol fermentation. In this study,the flocculant anchor gene flo1 and the cellulase gene cbh2 were fused and integrated to construct the recombinant expression vector pHBM368-PGK-flo1-cbh2. The linearized recombinant expression vector was electroporated into auxotrophic(Ura^-) S. cerevisiae INVSc cells.The cellulases were successfully expressed on the surface of S. cerevisiae cells,which was verified via functional screening and flow cytometry. The enzymatic activity of the whole cell enzyme was 66. 75 U/mL at50 ℃ for 1 h. In this study,we successfully constructed the whole cell S. cerevisiae cellulase,the results is useful for following whole-cell enzyme co-treating straw and its series of research.

关 键 词：酿酒酵母 细胞表面展示 纤维素酶 固定化酶 

分 类 号：Q939.97[生物学—微生物学]