Ito H, Hagiwara M, Takahashi K, Ichikizaki I
J Biochem. 1977 Sep;82(3):877-83. doi: 10.1093/oxfordjournals.jbchem.a131764.
Ribonuclease T1 [EC 3.1.4.8] was coupled to a water-insoluble cross-linked polyacrylamide (Enzacryl AH) by the acid azide method. The immobilized enzyme exhibited about 45% and 77% of the original activity toward yeast RNA and 2', 3-cyclic GMP, respectively, as substrates. Although the specific activity was lowered by the coupling, the immobilized enzyme was found to be far more stable to heat and extremes of PH than the native enzyme. The immobilized enzyme was active toward RNA even above pH 9 (at 37 degree C) or above 60 degree C (at pH 7.5), where the native enzyme was inactive. The immobilized enzyme retained much of its activity as assayed at 37 degree C after incubation in the range of pH 1 to 10 at 37 degree C, or after heating at 100 degree C (at pH 7.5) under conditions where the native enzyme was inactivated to a considerable extent. The enzyme derivative could be repeatedly recovered and reused without much loss of activity. The active site glutamic acid-58 in the immobilized enzyme appeared to be nearly as reactive with iodoacetate as that in the native enzyme.
核糖核酸酶T1 [EC 3.1.4.8] 通过酸叠氮法与水不溶性交联聚丙烯酰胺（Enzacryl AH）偶联。固定化酶分别以酵母RNA和2',3-环鸟苷酸为底物时，对它们的活性分别约为原来的45%和77%。虽然偶联后比活性降低，但发现固定化酶比天然酶对热和极端pH更为稳定。即使在pH 9以上（37℃）或60℃以上（pH 7.5），固定化酶对RNA仍有活性，而天然酶在此条件下无活性。在37℃下于pH 1至10范围内孵育后，或在天然酶在相当程度上失活的条件下于100℃（pH 7.5）加热后，固定化酶在37℃测定时仍保留其大部分活性。该酶衍生物可以反复回收和再利用，而活性没有太多损失。固定化酶中的活性位点谷氨酸-58与碘乙酸的反应性似乎与天然酶中的几乎一样。
