Zhang J, Corden J L
Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
J Biol Chem. 1991 Feb 5;266(4):2297-302.
The carboxyl-terminal domain (CTD) of the largest subunit of eukaryotic RNA polymerase II can be phosphorylated by a p34cdc2/CDC28-containing CTD kinase. Phosphorylated serine (or threonine) is located at positions 2 and 5 in the repetitive heptapeptide consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. We show here that phosphorylation of the mouse CTD retards its electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels in a way similar to that observed for the II0 form of the largest subunit of RNA polymerase II phosphorylated in vivo. At the maximum level of phosphorylation by CTD kinase in vitro, there are 15-20 phosphates evenly distributed among the 52 heptapeptide repeats that comprise the mouse CTD. Gel filtration chromatography and sucrose gradient ultracentrifugation analyses indicate that phosphorylation induces a dramatic conformational change in the CTD with the phosphorylated form adopting a far more extended structure than the unphosphorylated CTD.
真核生物RNA聚合酶II最大亚基的羧基末端结构域（CTD）可被含p34cdc2/CDC28的CTD激酶磷酸化。磷酸化的丝氨酸（或苏氨酸）位于重复七肽共有序列Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7的第2位和第5位。我们在此表明，小鼠CTD的磷酸化以类似于体内磷酸化的RNA聚合酶II最大亚基II0形式的方式，减缓其在十二烷基硫酸钠-聚丙烯酰胺凝胶中的电泳迁移率。在体外CTD激酶的最大磷酸化水平下，15-20个磷酸均匀分布在构成小鼠CTD的52个七肽重复序列中。凝胶过滤色谱和蔗糖梯度超速离心分析表明，磷酸化诱导CTD发生显著的构象变化，磷酸化形式比未磷酸化的CTD具有更伸展的结构。
