Flemming C, Gabert A, Gomoll M, Roth P
Acta Biol Med Ger. 1977;36(7-8):1007-18.
The kinetics of binding the enzymes glucose oxidase, trypsin, and peroxidase and also bovine gamma-globulin to the macroporous carriers dialdehyde cellulose, diazotized amino polystyrene, acrolein-acrylamide copolymer, and isothiocyanate groups carrying CPG-glass was studied. It was found empirically that the increase in protein content and--if there occurred no inactivation of enzyme during binding reaction--of enzymatic activity of the enzyme-carrier complex with the reaction time can be described by saturation curves (first order hyperbolas), which represent straight lines after suitable transformation. Thereby it is possible to calculate protein content or specific activity of the enzyme-carrier complex for any moment during binding reaction. From intercepts of the obtained straight lines with the ordinate and with the abscissa, respectively, one can determine the maximum binding capacity and the maximum specific activity and from their slope the reaction rate can be obtained. If there result no straight lines for plotting enzymatic activity of the enzyme-carrier complex against binding time after transformation, then this is an indication for partial inactivation of the free or already insolubilized enzyme. From kinetic measurements it is concluded that in all cases studied by use the preceding stage in chemical fixation of the enzyme to carrier is an adsorption equilibrium. Limitation of reaction rate by diffusion as in loading an ion exchanger could not be observed.
研究了葡萄糖氧化酶、胰蛋白酶、过氧化物酶以及牛γ-球蛋白与大孔载体二醛纤维素、重氮化氨基聚苯乙烯、丙烯醛-丙烯酰胺共聚物和带有异硫氰酸酯基团的CPG玻璃的结合动力学。通过实验发现，在结合反应过程中，如果酶没有失活，酶-载体复合物的蛋白质含量和酶活性随反应时间的增加可以用饱和曲线（一级双曲线）来描述，经过适当变换后这些曲线呈直线。由此可以计算结合反应过程中任何时刻酶-载体复合物的蛋白质含量或比活性。分别从所得直线与纵坐标和横坐标的截距可以确定最大结合容量和最大比活性，从其斜率可以得到反应速率。如果将酶-载体复合物的酶活性对结合时间作图，经过变换后得不到直线，那么这表明游离的或已经固定化的酶发生了部分失活。从动力学测量结果可以得出结论，在所研究的所有情况下，酶与载体化学固定的前一阶段都是吸附平衡。没有观察到像装填离子交换剂那样因扩散而限制反应速率的情况。
