山梨醇对黑曲霉源α-L-鼠李糖苷酶(r-Rha1)热稳定性的影响

Effect of Sorbitol on Thermal Stability ofα-L-Rhamnosidase(r-Rha1)from Aspergillus niger

提高酶热稳定性是工业酶属性改造的重要目标。山梨醇作为广泛应用于提高酶稳定性的外源添加剂,目前对其影响酶热稳定性的机制仍不明确。因此,作者以黑曲霉源α-L-鼠李糖苷酶(r-Rha1)为研究对象,通过热失活动力学、圆二色谱和荧光光谱的谱学分析,研究山梨醇对α-L-鼠李糖苷酶热稳定性的影响机制。结果表明,在60~70℃下,加入山梨醇可改善α-L-鼠李糖苷酶的热稳定性;在60、65、70℃条件下,加入1.2 mol/L山梨醇可分别将半衰期提高29、25、10倍;圆二色谱和荧光光谱结果表明山梨醇通过增强α-L-鼠李糖苷酶的表面疏水活性和结构刚性,从而提高其热稳定性。通过探究山梨醇对α-L-鼠李糖苷酶热稳定性的影响,进一步证明山梨醇提高酶热稳定性的普遍适用性,同时为后续α-L-鼠李糖苷酶的工业应用提供了参考。

The thermal stability of enzyme is an important target of industrial enzyme property modification.As an exogenous additive widely used to improve enzyme stability,the mechanism of sorbitol affecting enzyme thermal stability is still unclear.Therefore,the authors focused onα-L-rhamnosidase(r-Rha1)derived from Aspergillus niger and investigated the mechanism of sorbitol on the thermal stability ofα-L-rhamnosidase by thermal inactivation kinetics,circular dichroism and spectroscopic analysis of fluorescence mapping.Adding sorbitol improved the thermal stability ofα-L-rhamnosidase at 60~70℃,and 1.2 mol/L sorbitol increased the half-life ofα-L-rhamnosidase 29-fold,25-fold and 10-fold at 60,65℃and 70℃,respectively.The results of circular dichroism and fluorescence mapping plus indicated that sorbitol improved the thermal stability ofα-L-rhamnosidase by enhancing its surface hydrophobic activity and structural rigidity.The effect of sorbitol on the thermal stability ofα-L-rhamnosidase was investigated to further prove the general applicability of sorbitol in improving the thermal stability of the enzyme,while providing a reference for the subsequent industrial application ofα-L-rhamnosidase.