文献类型：期刊文献

中文题名：粪肠球菌β-N-乙酰氨基葡萄糖苷酶基因的克隆表达及酶学性质表征

英文题名：Cloning, Expression and Characterization of β-N-acetylglucosaminidase from Enterococcus faecalis

作者：陈宝莉[1];秦臻[1];赵黎明[1]

机构：[1]华东理工大学生物工程学院发酵工业分离提取技术研发中心生物反应器工程国家重点实验室,上海200237

年份：2019

卷号：9

期号：3

起止页码：296

中文期刊名：生物技术进展

外文期刊名：Current Biotechnology

收录：CSTPCD

基金：上海市曙光计划项目(15SG28);高等学校学科创新引智计划项目(B18022);中央高校基本科研业务费专项资金(22221818014)资助

语种：中文

中文关键词：β-N-乙酰氨基葡萄糖苷酶;粪肠球菌;克隆;表达;酶学性质

外文关键词：β-N-acetylglucosaminidase;Enterococcus faecalis;cloning;expression;characterization

摘要：N-乙酰氨基葡萄糖因其重要的功能活性而具有广阔的应用前景,β-N-乙酰氨基葡萄糖苷是酶法制备N-乙酰氨基葡萄糖领域一类重要的糖苷水解酶。从粪肠球菌(Enterococcus faecalis)基因组中克隆得到一个GH20家族β-N-乙酰氨基葡萄糖苷酶基因(EfNagase),并在大肠杆菌中进行了异源表达,研究了重组酶的酶学性质。序列分析发现EfNagase与已报道的Streptococcus pneumoniae TIGR4来源的β-N-乙酰氨基葡萄糖苷酶序列相似度最高,为52.08%。对其酶学性质研究发现纯化后的EfNagase比活力为3 606.10 U/mg,在温度50℃、pH 6.0的条件下活性最高,且在温度低于50℃、pH 3.5～11.5的范围内展现出较高的稳定性(残余酶活>90%)。β-N-乙酰氨基葡萄糖苷酶是酶法制备N-乙酰氨基葡萄糖中重要的一类糖苷水解酶。研究发现EfNagase具有高酶活、高热稳定性和严格的底物特异性,在酶法制备N-乙酰氨基葡萄糖领域具有良好的应用潜力。
N-acetylglucosamine has broad application prospects due to its important functional activity.β-N-acetylglucosaminide is an important glycoside hydrolase in the field of enzymatic preparation of N-acetylglucosamine. A β-N-acetylaminoglycosidase gene from Enterococcus faecalis was cloned and heterologously expressed in Escherichia coli BL21. EfNagase showed the highest sequence similarity with Streptococcus pneumoniae TIGR4 β-N-acetylglucosaminidase (52.08%). The purified recombinant EfNagase displayed its activity on pNP-GlcNAc with specific activity 3 606.10 U/mg. It showed optimal activities at 50℃ and pH 6.0. It exhibited ultra-high stability (residual enzyme activity >90%) at temperatures below 50°C and pH 3.5~11.5.β-N-acetylglucosaminidase is an important class of glycoside hydrolase in the preparation of N-acetylglucosamine by enzymatic method. This study found that EfNagase had high enzymatic activity, high thermostability and strict substrate specificity, which has the potential to be used for the efficient preparation of N-acetylglucosamine.