采用复合保护剂提高重组普鲁兰酶稳定性

Enhancement of stability of recombinant Pullulanase by combination additive

通过向重组普鲁兰酶(PUL)酶液中添加保护剂和防腐剂以提高其热稳定性及储存稳定性,在不同温度下研究了复合保护剂对酶液的热稳定性影响,并研究了保护剂和防腐剂在常温保藏下对酶液贮存稳定性的影响。结果表明,明胶、甘油、Tween20和Ca2+四种保护剂能显著提高PUL的热稳定性,并通过正交试验得到了效果最佳的复合保护剂配方,即Tween20 2 m L/L,明胶6 g/L,甘油5%,Ca2+0.1 mol/L。在60℃下,添加复合保护剂的PUL酶液保温1 h后的酶活保留率为96.07%,较对照高出85.29%;在55℃,通过添加复合保护剂,半衰期较对照提高了3.27倍。最终在常温保藏90 d后,含有复合保护剂和防腐剂(1 g/L山梨酸钾和1 g/L苯甲酸钠)的PUL酶活保留率高达85.25%,达到了工业保藏的需求。

To enhance the thermostability and storage stability of recombinant pullulanase( PUL),protective additives and preservatives were added into the preparation of PUL,and the effects of combination additive on the thermostability of PUL at different temperatures were studied. Then the effects of combination additive and preservative on the storage stability of PUL at 25 ℃ were also investigated. The results indicated that gelatin,glycerin,Tween 20 and Ca2 +could significantly improve the thermostability of PUL. The optimal combination additive( Tween 20 2 m L / L,gelatin 6 g / L,glycerin 5%,Ca2 +0. 1 mol / L) was obtained using the orthogonal experimental design. The residual activity of PUL with combination additive maintained 96. 07% after incubating 1 h at 60 ℃,which was 85. 29% higher than that of control without protective additive. Furthermore,the half-life of PUL with combination additive was increased by 3. 27 times compared to that of control at 55 ℃. Finally,the residual activity of PUL containing combination additive and preservatives( 0. 1% potassium sorbate,0. 1% sodium benzoate) reached 85. 25% after storing at room temperature for 90 days,and thus met the requirements of industrialization.