Bispora sp. MEY-1来源的嗜酸海藻糖酶TreA酶学性质研究

Enzymatic Properties of an Acidic Trehalase TreA from Bispora sp.MEY-1

酸性海藻糖酶作为生物催化剂在生物工业中广泛应用,挖掘酶学性质优良的酸性海藻糖基因具有重要意义。从Bispora sp.MEY-1中成功克隆出一个酸性海藻糖酶基因TreA,并在毕赤酵母GS115中实现高效表达。对纯化后的酸性海藻糖酶TreA进行酶学性质鉴定,其最适pH为4.0,在pH 2.2~5.0范围内,该酶能够维持60%以上的酶活力。TreA在偏酸条件下稳定效果比较好,在pH 1.0~4.0条件下处理1 h,可以剩余88%以上的酶活力。TreA的最适温度为60℃,在40~70℃之间都可以维持50%以上的酶活力,属于中高温酶。重组海藻糖酶TreA的比活为1913 U·mg^(-1),动力学参数K_m和V_(max)分别为0.67 mg·mL^(-1)和119μmol·min^(-1)·mg^(-1)。同时,还评估了不同金属离子或化学试剂对纯海藻糖酶TreA的酶活性影响及其蛋白酶抗性。极大丰富了海藻糖酶基因资源,促进了其在生物工业中的应用。

As a biocatalystase,acid trehalase is widely used in biological industry.It is very important to exploit new acid trehalase with excellent enzymatic properties.In this study,an acid trehalase gene(TreA)from Bispora sp.MEY-1 was successfully cloned and expressed in Pichia pastoris GS115.The enzymatic properties of purified trehalase TreA were identified.The optimum pH was 4.0,and the relative activity maintained over 60%under the range of pH 2.2~5.0.The relative activity of purified enzyme was more than 88%after treated for 1 h under pH 1.0~4.0,which exhibited the good acid stability.The optimum temperature of purified enzyme was 60℃,which was enzyme with medium-high temperature resistance.The specific activity of purified enzyme was 1913 U·mg^(-1),and its K m and V max were 0.67 mg·mL^(-1)and 119μmol·min^(-1)·mg^(-1),respectively.Moreover,the effects of different mental ions or chemical reagents,and the protease resistances on the purified enzyme activity were also evaluated.