超嗜热细菌Thermotoga maritima酯酶Tm1350克隆、表达及其酶学性质研究  

Cloning,expression and characterization of the esterase Tm1350 from hyperthermophilic bacteria Thermotoga maritima

作　　者：魏涛[1] 杜聪聪[1] 毛多斌[1] 马歌丽[1] 

机构地区：[1]郑州轻工业学院食品与生物工程学院,河南郑州450002

出　　处：《食品工业科技》2015年第9期179-183,共5页Science and Technology of Food Industry

基　　金：河南省重点科技攻关项目(132102120197)

摘　　要：以超嗜热细菌Thermotoga maritima基因组DNA为模板,通过PCR法扩增酯酶Tm1350基因,构建重组质粒p ET15b-Tm1350,在大肠杆菌Escherichia coli BL21(DE3)中实现表达,并采用p-NPA方法测定其酶活性。研究结果表明,酯酶Tm1350可以水解不同碳链长度(10C)的对硝基苯酚酯,其中对硝基苯酚戊酸酯(p NP-C5)是该酶最适合底物;该酶最适反应温度和p H分别为70℃和6.5,90℃处理5h,仍有50%以上的酶活力,具有较强的热稳定性。酯酶Tm1350对金属离子、抑制剂、变性剂和有机溶剂具有较强的抗性。The esterase gene Tm1350 from Thermotoga maritima was amplified by PCR method and recombinant plasmid p ET15b- Tm1350 was constructed to express the enzyme in Escherichia coli BL21（ DE3）.Esterase activity was determined spectrophotometrically by the p- NPA method using p- nitrophenyl esters of various chain lengths as the substrates.The enzyme could hydrolyze the substrates of p NP- esters with acyl chains of different lengths and exhibited the highest activity for p NP- C5 among the substrates tested. Tm1350 displayed optimal activity at70℃ and 6.5.It maintained above 50% activity after 5h incubation at 90℃,which indicated that the enzyme showed strong thermal stability. The recombinant esterase Tm1350 was found to have strong resistances to metal ions,inhibitor,denaturant and organic solvents.

关 键 词：THERMOTOGA maritima 酯酶 克隆表达 酶学性质 

分 类 号：TS201.3[轻工技术与工程—食品科学]