木聚糖酶XynB-E18的重组表达纯化及酶学性质测定  

Recombinant Expression Purification of Xylanase XynB-E18 and Determination of the Enzymological Properties

作　　者：宋迎 鲁芳[1] 严孝金[1] 张蓓[1] 刘永[1] 张立[1] 孟昆[2] 杨培龙[2] 姚斌 高伟[1] SONG Ying;LU Fang;YAN Xiao-jin;ZHANG Bei;LIU Yong;ZHANG Li;MENG Kun;YANG Bei-long;YAO Bin;GAO Wei(School of Science/Beijing Forestry University,Beijing 100083,China;Chinese Academy of Agricultural Sciences,Beijing 100081,China)

机构地区：[1]北京林业大学理学院,北京100083 [2]中国农业科学院,北京100081

出　　处：《山东农业大学学报（自然科学版）》2018年第2期320-325,共6页Journal of Shandong Agricultural University：Natural Science Edition

基　　金：国家自然科学基金(31070651);中央高校基本科研基金(2015ZCQ-LY-02)

摘　　要：为了测定木聚糖酶Xyn B-E18的活性以及酶学特征,采用大肠杆菌原核表达系统对来源于青贮饲料菌群的木聚糖酶Xyn B-E18进行重组表达纯化。在16℃180 r·min-1条件下表达,并通过Ni亲和层析和分子排阻层析的方法进行纯化,用DNS显色法测定其酶学性质。得到的重组蛋白能在大肠杆菌中获得高效表达,纯化后的蛋白溶液纯度达90%以上,蛋白分子量约为38k Da,酶活力可达1782 U,最适反应p H为6.5,在中性偏碱性条件下酶的稳定性较好,最适反应温度为50℃,在50℃以下酶的热稳定性保持较好。实验结果表明Xyn B-E18具有良好的稳定性,可具有更大应用潜力。In order to determine the activity and enzymatic characteristics of xylanase XynB-E18,the recombinant expression and purification of xylanase XynB-E18 from silage bacteria was carried out by using E.coli prokaryotic expression system.It was expressed at 16℃at 180 r?min-1 and purified by Ni affinity chromatography and molecular exclusion chromatography.The enzymatic properties were determined by DNS colorimetric method.The recombinant XynB-E18 protein can be expressed efficiently in Escherichia coli,and its purity was more than 90%;The molecular weight of the protein was about 38 kDa.The enzyme activity can reach 1782 U,and the optimum reaction pH was 6.5;The stability of the enzyme was better under neutral and alkaline conditions,the optimum reaction temperature was 50℃,and the thermal stability presented a good performance below 50°?.The experimental results show that XynB-E18 has good stability and has greater application potential.

关 键 词：XynB-E18 重组表达 分离纯化 酶学特性 

分 类 号：Q6[生物学—生物物理学]