抗坏血酸对酵母蔗糖酶的激活动力学研究

采用甲苯自溶法从鲜酵母中提取了蔗糖酶，并用乙醇分级及ＤＥＡＥ－纤维素柱层析进行了纯化，用ＰＡＧ凝胶电泳作了纯度鉴定，在ｐＨ５．０，３０℃条件下进行了酶反应，用双倒数作图法测出其Ｋｍ＝２．１×１０－２ｍｏｌ／Ｌ，Ｖｍａｘ＝０．２６（每分钟的光密度值）．在此系统中，加入不同浓度的抗坏血酸（Ｖｉｔ．Ｃ），发现其具有激活作用并存在量效关系．双倒数作图显示：酶的表观Ｖｍａｘ（Ｖｐ）随抗坏血酸浓度的增加而增大，但其表观Ｋｍ（Ｋｐ）不变（Ｋｐ＝Ｋｍ）．经实验结果分析，推论出抗坏血酸激活作用的酶促反应方程式，并推导出反应速度公式

The Kinetics of Activation of Yeast Invertase by Ascorbic Acid

Invertase was extracted from fresh yeast with adding methylbenzene and gradationally purified with different ethanol concentrations which were 32% and 47.5%, respectively.Purity appeared homogeneous, which was identified by PAGE.The enzyme reaction was performed at 30℃ and pH 5.0 (maintained with 0.2 mol/L acetate buffer). K m(10 -2 mol/L) and V max [0.26(optical density perminute)] had been obtained by a set of double reciprocal plot, respectively.In this reaction, when varying concentration of sucrose was added at changing fixed levels of ascorbic acid, the concentrations of ascorbic acid were: 0 mmol/L, 2 mmol/L,4 mmol/L,6 mmol/L,8 mmol/L and 10 mmol/L.It was found that ascorbic acid functioned as activator and had stoichiometric relation to its effect.A set of double reciprocal plot showed that apparent V max ( V p) increased with an increase in ascorbic acid concentration, but apparent K m( K p) was unchanged ( K p= K m).The enzymatic reaction equation of ascorbic acid activation has been deducted and then rate formula of the reaction derived by analyzing of experiment results.