两种化学修饰剂对小麦酯酶活性及稳定性的影响

Two Chemical Modifiers Affect Wheat Esterase Activity and Stability

目的:用右旋糖酐和单甲氧基聚乙二醇(mPEG)化学修饰纯化后的小麦酯酶,并考察修饰酶的酶学性质。方法:分别采用高碘酸钠和三聚氯氰活化右旋糖酐和mPEG,得到修饰的小麦酯酶,通过红外光谱表征修饰酶基团变化,测试修饰酶的反应动力学参数,并考察温度及pH值对修饰酶活力的影响。结果:修饰后的酶与纯化酶相比,修饰酶对底物乙酸-1-萘酯的亲和力增强,并保留了大部分酶的活力,右旋糖酐修饰酶活保持率在67.1%,mPEG修饰酶的酶活保持率达到87.7%。另外,右旋糖酐修饰酶的最适温度和pH值分别为35℃和6.4,K m和V max分别为2.00mmol/L和2000 g/min,而mPEG修饰酶的最适温度和pH值为35℃和6.8,K m和V max为1.25mmol/L和2500 g/min。两种修饰酶在耐热性、耐酸碱性等方面都优于纯化酶,mPEG修饰酶的最适pH值发生改变。结论:修饰酶较纯化酶更稳定,具有一定的实用意义。

Objective： To modify purified wheat esterase with dextran and methoxy polyethyleneglycol （mPEG）, and characterizethe modified enzymes. Methods： Dextran and mPEG were activated by sodium periodate and trimer cyanuric chloride, respectively,before being used to prepare the modified wheat esterases. Changes in the active groups in the modified enzyme moleculeswere described through infrared spectroscopy, and their kinetic parameters and the effects of temperature and pH on the modifiedenzyme activities were investigated. Results： In comparison with the native enzyme, the modified enzyme had better affinity for the substratealpha-naphthyl acetate, and retained the majority of enzyme activity. The residual activity of dextran-modified enzyme was 67.1%,and the mPEG modified enzyme retained 87.7% of the original activity. In addition, thermal stability and acid-base stability of the twomodified enzymes were better than those of the native one, and the optimal pH of the mPEG-modified enzyme was changed. Conclusion：The modified enzymes have more stability than the native enzyme and thus are of important practical significance.