Götz C, Koenig M G, Issinger O G, Montenarh M
Department of Medical Biochemistry, University of the Saarland, Homburg, Germany.
Eur J Biochem. 1995 Oct 1;233(1):327-34. doi: 10.1111/j.1432-1033.1995.327_1.x.
The simian virus 40 (SV40) large T antigen is a multifunctional protein involved in SV40 cell transformation and lytic virus infection. Some of its activities are regulated by interaction with cellular proteins and/or by phosphorylation of T antigen by various protein kinases. In this study, we show that immuno-purified T antigen from SV40-transformed cells and from baculovirus-infected insect cells is tightly associated with a protein kinase that phosphorylates T antigen in vitro. In the presence of heparin or a peptide resembling a protein kinase CK2 recognition site, the phosphorylation of T antigen by the associated kinase is reduced whereas a p34cdc2-kinase-specific peptide has no influence. In addition, the T-antigen-associated protein kinase can use GTP and ATP as phosphate donors. These properties together with the observation that immunopurified T antigen can be phosphorylated by the addition of protein kinase CK2 suggest that at least one of the T-antigen-associated protein kinases is CK2 or a protein-kinase-CK2-related enzyme. The association of recombinant CK2 with T antigen was strongly confirmed by in vitro binding studies. Experiments with temperature-sensitive SV40-transformed cells provide evidence for a close correlation between cell transformation and phosphorylation of T antigen by the associated protein kinase.
猿猴病毒40（SV40）大T抗原是一种多功能蛋白，参与SV40细胞转化和裂解性病毒感染。其一些活性通过与细胞蛋白相互作用和/或通过各种蛋白激酶对T抗原的磷酸化来调节。在本研究中，我们表明，从SV40转化细胞和杆状病毒感染的昆虫细胞中免疫纯化的T抗原与一种在体外使T抗原磷酸化的蛋白激酶紧密相关。在存在肝素或类似蛋白激酶CK2识别位点的肽的情况下，相关激酶对T抗原的磷酸化作用降低，而p34cdc2激酶特异性肽则没有影响。此外，与T抗原相关的蛋白激酶可以使用GTP和ATP作为磷酸供体。这些特性以及免疫纯化的T抗原可通过添加蛋白激酶CK2进行磷酸化这一观察结果表明，与T抗原相关的蛋白激酶中至少有一种是CK2或与蛋白激酶CK2相关的酶。体外结合研究有力地证实了重组CK2与T抗原的结合。对温度敏感的SV40转化细胞进行的实验为细胞转化与相关蛋白激酶对T抗原的磷酸化之间的密切相关性提供了证据。
