Renart M F, Sastre L, Díaz V, Sebastián J
Mol Cell Biochem. 1985 Feb;66(1):21-9. doi: 10.1007/BF00231819.
A purification procedure to obtain RNA polymerases I (or A) and II (or B) from Dictyostelium discoideum amoeba has been developed. The enzymes were solubilized from purified nuclei and separated by DEAE-Sephadex chromatography. RNA polymerases I and II were further purified by a second chromatography on DEAE-Sephadex followed by chromatographies on phosphocellulose and heparin-sepharose. The specific activities of purified RNA polymerases I and II are 92 units/mg protein and 70 units/mg protein, respectively. The subunit structure of both RNA polymerases were analyzed by polyacrylamide gel electrophoresis under denaturing conditions after glycerol gradient centrifugation of the enzymes. The putative subunits of RNA polymerase I have molecular weights of 180 000, 125 000, 43 000, 40,000, 34 000, 31 000, 25 000, 19 000, 17 000 and 14 000. The putative subunits of RNA polymerase II have molecular weights of 200 000 (170 000), 130 000, 33 000, 25 000, 19 000, 17 000, 15 000, 13 000. There are three polypeptides with common molecular weight in Dictyostelium RNA polymerases I and II. The subunit of 25 000 daltons of both enzymes has common immunological determinants with RNA polymerase II from crustacean Artemia.
已开发出一种从盘基网柄菌变形虫中获取RNA聚合酶I（或A）和II（或B）的纯化程序。这些酶从纯化的细胞核中溶解出来，并通过DEAE-葡聚糖凝胶色谱法进行分离。RNA聚合酶I和II通过在DEAE-葡聚糖凝胶上进行第二次色谱，随后在磷酸纤维素和肝素-琼脂糖上进行色谱进一步纯化。纯化后的RNA聚合酶I和II的比活性分别为92单位/毫克蛋白质和70单位/毫克蛋白质。在对酶进行甘油梯度离心后，通过变性条件下的聚丙烯酰胺凝胶电泳分析了两种RNA聚合酶的亚基结构。RNA聚合酶I的推定亚基分子量分别为180000、125000、43000、40000、34000、3100,00、25000、19000、17000和14000。RNA聚合酶II的推定亚基分子量分别为200000（170000）、130000、33000、25000、19000、17000、15000、13000。盘基网柄菌RNA聚合酶I和II中有三种多肽分子量相同。两种酶25000道尔顿的亚基与甲壳动物卤虫的RNA聚合酶II具有共同的免疫决定簇。
