溶藻弧菌中酯酶基因的克隆表达及其酶学性质研究  

Cloning,Expression and Characterization of the Esterase Gene estZ from Vibrio alginolyticus

作　　者：赵书梅[1] 王霖慧 唐嘉琦 赵静宜[1] 

机构地区：[1]中国石油大学(华东),青岛266580

出　　处：《生物技术通报》2017年第6期190-196,共7页Biotechnology Bulletin

基　　金：山东省中青年科学家科研奖励基金(ZR2016CB11);中央高校基本科研业务费专项资金(16CX02043A)

摘　　要：旨在克隆溶藻弧菌中的酯酶基因,并在大肠杆菌中异源表达,研究酯酶酶学性质。从溶藻弧菌(Vibrio alginolyticus)7S-1的基因组DNA中克隆得到酯酶基因est Z,将该基因连接入表达载体p ET28a,并在Escherichia coli BL21(DE3)中表达,对纯化后的酯酶蛋白进行酶学性质分析。结果显示,表达菌株E.coli BL21-28a-est Z可高表达具有活性的酯酶,在18℃添加0.4 mmol/L IPTG的LB培养基中诱导22 h,酯酶的比酶活可达5.42 U/μg;重组表达的酯酶Est Z最适反应温度为25℃,最适pH为9.0。10 mmol/L Na+和Mg^(2+)对其有较好的激活作用;Cu^(2+)、Ni^(2+)、Co^(2+)对该酶活性有较大抑制。该酶对多种有机溶剂及表面活性剂具有一定的耐受性,同时具有较高的耐盐性。对溶藻弧菌中的酯酶进行研究为该菌更好的应用于生物脱墨技术奠定了理论基础。This work aimed to investigate the esterase gene estZ of Vibrio alginolyticus 7S-1,which was expressed in Escherichia coliand characterized. The estZ gene was obtained from the genomic DNA of V. alginolyticus 7S-1 by gene cloning,then ligated to the vectorp ET28 a,and expressed in E. coli BL21（DE3）. E. coli BL21-28a-est Z expressed the est Z gene with esterase activity. The specific activity ofthe purified protein Est Z reached 5.42 U/μg when the E. coli cells were induced by 0.4 mmol/L IPTG and grew at 18℃ for 22 hours. The optimaltemperature and p H of the Est Z protein were 25℃ and 9.0,respectively. Est Z was activated by 10 mmol/L Na＋ and Mg^（2＋）,but highly inhibitedby Cu^（2＋）,Ni^（2＋） and Co^（2＋）. In addition,Est Z is resistant to some organic solvents,surfactants and salt. This study would provide some insights of V. alginolyticus in biological deinking industry.

关 键 词：溶藻弧菌 酯酶 酶学性质 

分 类 号：Q78[生物学—分子生物学] Q936