来源于海洋细菌Altererythrobacter luteolus SW109~T的新型酯酶E29的克隆表达及其酶学性质  

Cloning, expression and characterization of a novel esterase(E29) from a marine bacterium Altererythrobacter luteolus SW109~T

作　　者：李正阳[1] 戎振 王昭[1,2] 霍颖异[1] 孟凡旭[1] 王春生[1] 崔恒林[2] 许学伟[1] 

机构地区：[1]国家海洋局第二海洋研究所国家海洋局海洋生态系统与生物地球化学重点实验室,浙江杭州310012 [2]江苏大学食品与生物工程学院,江苏镇江212013

出　　处：《微生物学通报》2016年第5期1051-1059,共9页Microbiology China

基　　金：国家重点基础研究发展规划项目(973计划)(No.2014CB441503);国家自然科学基金项目(No.41506183)~~

摘　　要：【目的】克隆表达一个来源于海洋细菌的酯酶E29,并研究其酶学性质。【方法】从细菌Altererythrobacter luteolus SW109T中筛选并扩增出一个酯酶基因,将其克隆至p SMT3载体上,并将重组质粒转化至大肠杆菌BL21(DE3)中进行异源表达,分析表达产物的酶学性质。【结果】氨基酸序列分析结果表明,酯酶E29属于脂类水解酶第二家族(Family II)。酶学性质分析结果显示,酯酶E29的最适反应底物为对硝基苯酚丁酸酯,最适反应温度为45°C,最适反应pH为8.5;10 mmol/L的Co^(2+)和Mn^(2+)及15%的异丙醇和乙腈能强烈抑制酯酶E29的活性,1%的SDS能使酶失活,甘油的存在能促进酶活性。【结论】酯酶E29是一个海洋来源的新型酯酶,其具有较高的酶活力值、较宽的底物谱以及对部分有机溶剂和金属离子较好的耐受性,在工业方面具有潜在的应用价值。[Objective] We cloned, expressed and characterized a novel esterase E29 from a marine bacterium. [Methods[ An assumed esterase gene, which was predicted via the genome ofAltererythrobacter luteolus SW109T and amplified by PCR, was cloned into expression vector pSMT3. The recombinant plasmid was transformed into Escherichia coli BL21（DE3）. Subsequently, E29 was obtained via heterogenetic expression and characterized. [Resultsl The amino acids sequence analysis revealed E29 represents a new member of Family II of lipolytic enzyme. According to biochemical characterization, the maximum hydrolysis activity was obtained using p-nitrophenyl butyrate as substrate, at 45 ℃ and pH 8.5. The activity of E29 decreased in solution containing 10 mmol/L Co2＋ or Mn2＋, or 15% isopropanol or acetonitrile. The esterase was inactivated in 1% SDS solution. The addition of glycerol can promote the activity of E29 strikingly. [Conclusion] E29 is a novel marine esterase. Due to its high enzyme activity, wide substrates selectivity as well as tolerance of some organic solvents and metal irons, E29 has potential application in industry.

关 键 词：海洋细菌 酯酶 酶学性质 克隆表达 

分 类 号：Q936[生物学—微生物学]