Koroleva E I, Bulgadaeva S A, Grigorovich Iu A
Biokhimiia. 1979 Mar;44(3):555-63.
Beef heart cAMP phosphodiesterase (EC 3.1.4.17) was isolated and partially purified using fractionation by ammonium sulfate and gel filtration on the columns with Sephadex G-200 and Sepharose 6B. This method allowed to preserve the enzyme binding to the low-molecular weight thermostable protein regulator of the phosphodiesterase activity. The enzyme preparation was purified 130--180-fold as compared to the original homogenate. The pH-dependence of the enzyme activity in the imidazole and tris -- buffers for the fraction with maximal activity was carried out. The kinetic analysis of this fraction revealed an abnormal kinetic behaviour with two Km values. The enzyme is represented by four forms differing in their molecular weights and possessing different capacity for activation by Ca2+ and protein regulator. No activation was observed in the forms with higher molecular weights, whereas the activity of the forms with lower molecular weights depended on the presence of Ca2+ and protein regulator. It is assumed that some of the above-described forms are capable of interconversions.
利用硫酸铵分级分离以及在装有葡聚糖G - 200和琼脂糖6B的柱上进行凝胶过滤，分离并部分纯化了牛心cAMP磷酸二酯酶（EC 3.1.4.17）。该方法能够保留酶与磷酸二酯酶活性的低分子量热稳定蛋白调节剂的结合。与原始匀浆相比，酶制剂纯化了130 - 180倍。对具有最大活性的级分在咪唑和tris缓冲液中进行了酶活性的pH依赖性研究。对该级分的动力学分析揭示了具有两个Km值的异常动力学行为。该酶由四种分子量不同的形式组成，并且具有不同的被Ca2 +和蛋白调节剂激活的能力。分子量较高的形式未观察到激活现象，而分子量较低的形式的活性取决于Ca2 +和蛋白调节剂的存在。据推测，上述一些形式能够相互转化。
