Sulfolobus acidocaldarius ATCC 33909麦芽寡糖基海藻糖水解酶突变株的酶学性质及其制备海藻糖的条件优化

Enzymatic properties of maltooligosyltrehalose trehalohydrolase mutant strain from Sulfolobus acidocaldarius ATCC 33909 and optimization of trehalose preparation

来源于古细菌嗜酸热硫矿硫化叶菌(Sulfolobus acidocaldarius ATCC 33909)的麦芽寡糖基海藻糖水解酶(maltooligosyltrehalose trehalohydrolase,MTHase)是双酶法生产海藻糖的关键酶。对1株蛋白表达量提高1.9倍的突变株L202P/L218D/Y323G进行酶学性质方面的测定和海藻糖转化。对突变株L202P/L218D/Y323G Sa MTHase进行最适温度、最适pH值、pH稳定性和60℃半衰期的测定,发现该突变株的酶学性质未发生较大变化。将麦芽四糖基海藻糖为底物,测定突变株L202P/L218D/Y323G Sa MTHase酶动力学相关参数。将麦芽糊精(DE值5~7)作为底物,利用嗜酸热硫矿硫化叶菌的麦芽寡糖基海藻糖合成酶(maltooligosyl trehalose synthase,MTSase)与突变株L202P/L218D/Y323G Sa MTHase以不同比例作用于底物生产海藻糖,海藻糖的转化率最高达到76.0%。

Maltooligosyltrehalose trehalohydrolase(MTHase),which is derived from the Sulfolobus acidocaldarius ATCC 33909,is a key enzyme in the couple enzymatic production of trehalose.The mutant strain L202P/L218D/Y323G,which has a 1.9-fold increase in protein expression,was used for the determination of enzymatic properties and trehalose conversion.The optimum temperature,optimum pH,pH stability and 60℃half-life of mutant strain L202P/L218D/Y323G SaMTHase were determined.The results showed that there was no significant change in the enzymatic properties of the mutant strain.The kinetic parameters of the mutant strain L202P/L218D/Y323G SaMTHase were determined using maltotetraosyl trehalose as substrate.Using Maltodextrin(DE 5-7)as substrate,maltooligosyl trehalose synthase(MTSase)was mixed with the mutant L202P/L218D/Y323G SaMTHase at different ratios.The trehalose conversion results showed that trehalose conversion ratio was up to 76.0%.