壳聚糖固定化海藻糖合酶

利用壳聚糖作为载体,将大肠杆菌BL21异源表达且纯化后的海藻糖合酶(TreS),采用吸附法制备成固定化酶。固定化单因素实验结果表明,最适海藻糖合酶与壳聚糖的加入量为32.0 mg/g,最适吸附时间为2.5 h。固定化酶最适反应温度为40 ℃,最适反应pH为6.0,酶活回收率为40.17%,重复使用9次后,固定化酶酶活残留率为64.64%,重复使用性良好。在4~60 ℃范围内放置20 min后,固定化酶的相对酶活均高于87%,与游离酶相比温度稳定性没有明显变化。在pH3.5~8.5范围内放置20 min后,固定化酶的相对酶活均高于60%,酸碱稳定性优于游离酶。反应进程试验结果表明,2 h时海藻糖得率达最大,为61.4%。

Immobilization of Trehalose Synthase with Chitosan as the Carrier

In this research, trehalose synthase (TreS) was expressed using Escherichia coli BL21.Then, the purified enzyme was immobilized by chitosan according to adsorption method. After single-factor experiments, the optimal amount of the immobilized enzyme was 32.0 mg per gram of chitosan, the optimum time of binding was 2.5 hours. The optimum reaction temperature and pH of the immobilized enzyme were 40℃ and 6.0, respectively. The highest activity recovery of the immobilized enzyme was 40.17%. The rate of residual enzyme activity of the immobilized enzyme was 64.64% after consecutive 9 cycles, which had reusability a good. The thermal stability of the immobilized enzyme was not significantly different compared with the free enzyme, and the relative activity maintained 87% after preincubation of the immobilized enzyme at different temperatures (4~60℃) for 20 min. However, the pH stability of the immobilized enzyme was superior, and the relative activity still retained 60% after preincubation of the immobilized enzyme in various pH values (3.5~8.5) for 20 min. The results of reaction processes indicated that the maximum trehalose yield of this immobilized enzyme was 61.4% at the point of 2 hours.