Vaisius A C, Horgen P A
Biochemistry. 1979 Mar 6;18(5):795-803. doi: 10.1021/bi00572a009.
The DNA-dependent RNA polymerases II or B (ribonucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) from the mushroom Agaricus bisporus has been purified to apparent homogeneity. The purification procedures involve precipitation with polyethylenimine, selective elution of RNA polymerase II from the polyethylenimine precipitate, ammonium sulfate fractionation, DEAE-cellulose chromatography, CM-cellulose chromatography, and exclusion chromatography on Bio-Gel A-1.5M. With this procedure 11 mg of RNA polymerase II is recovered from 1.5 kg of mushroom tissue. RNA polymerase II from Agaricus bisporus has 12 subunits with the following molecular weights: 182,000, 140,000, 89,000, 69,000, 53,000, 41,000, 37,000, 31,000, 29,000, 25,000, 19,000, and 16,500. Purified RNA polymerase II from Agaricus bisporous was half-maximally inhibited by the mushroom toxin alpha-amanitin at a concentration of 6.5 microgram/mL (7 X 10(-6) M), which is 650-fold more resistant than mammalian RNA polymerases II. The apparent Ki for the alpha-amanitin-RNA polymerase complex was estimated to be 12 X 10(-6) M. The activity of purified RNA polymerase II from the mushroom was quite typical of other eukaryotic RNA polymerase II with regard to template preference, salt optima, and divalent metal cation optima.
双孢蘑菇的DNA依赖性RNA聚合酶II或B（核糖核苷三磷酸：RNA核苷酸转移酶，EC 2.7.7.6）已被纯化至表观均一。纯化步骤包括用聚乙烯亚胺沉淀、从聚乙烯亚胺沉淀中选择性洗脱RNA聚合酶II、硫酸铵分级分离、DEAE-纤维素色谱、CM-纤维素色谱以及在Bio-Gel A-1.5M上进行排阻色谱。通过该方法，从1.5千克蘑菇组织中回收了11毫克RNA聚合酶II。双孢蘑菇的RNA聚合酶II有12个亚基，分子量如下：182,000、140,000、89,000、69,000、53,000、41,000、37,000、31,000、29,000、25,000、19,000和16,500。双孢蘑菇纯化的RNA聚合酶II在浓度为6.5微克/毫升（7×10⁻⁶M）的蘑菇毒素α-鹅膏蕈碱作用下受到半数抑制，这比哺乳动物RNA聚合酶II的抗性高650倍。α-鹅膏蕈碱-RNA聚合酶复合物的表观Ki估计为12×10⁻⁶M。就模板偏好、最适盐浓度和二价金属阳离子最适浓度而言，蘑菇纯化的RNA聚合酶II的活性是其他真核RNA聚合酶II的典型特征。
