Defining the substrate specificity of cdk4 kinase-cyclin D1 complex
Defining the substrate specificity of cdk4 kinase-cyclin D1 complex
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DOI：
10.1093/carcin/20.2.193
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发表时间：
1999-02-01
期刊：
影响因子：
4.7
通讯作者：
Hoess, RH
中科院分区：
文献类型：
--
作者：
Grafstrom, RH;Pan, WJ;Hoess, RH
cdk 4激酶-细胞周期蛋白D1复合物（cdk 4/D1）不能同等地磷酸化视网膜母细胞瘤蛋白（Rb）内的所有位点。比较Rb 15 kDa C结构域内的5个磷酸化位点表明，Ser 795是cdk 4/D1磷酸化的优选位点。已经进行了一系列实验以确定该位点指导优先磷酸化的性质。对于cdk 4/D1，在磷酸化的丝氨酸/苏氨酸的C末端的第三位的优选氨基酸是精氨酸，其他氨基酸的取代，包括对赖氨酸的保守改变，对磷酸化速率有显着影响。该信息已用于突变Rb中不太有利的位点，将它们转化为现在优先被cdk 4/D1磷酸化的位点，相关口袋蛋白p107中Ser 842的保守位点也优先被cdk 4/D1磷酸化。尽管Rb和p107在序列上显著不同，但Rb Ser 795位点可以取代p107 Ser 842位点而不影响磷酸化速率，这些结果表明，虽然特异性的决定因素存在于磷酸化位点周围的序列中，但该位点的结构背景也是特异性的关键参数。
cdk4 kinase-cyclin D1 complex (cdk4/D1) does not phosphorylate all of the sites within retinoblastoma protein (Rb) equally. Comparison of five phosphorylation sites within the 15 kDa C domain of Rb indicates that Ser795 is the preferred site of phosphorylation by cdk4/D1, A series of experiments has been performed to determine the properties of this site that direct preferential phosphorylation, For cdk4/D1, the preferred amino acid at the third position C-terminal to the phosphorylated serine/threonine is arginine, Substitution of other amino acids, including a conservative change to lysine, has dramatic effects on the rates of phosphorylation, This information has been used to mutate less favorable sites in Rb, converting them to sites that are now preferentially phosphorylated by cdk4/D1, A conserved site at Ser842 in the related pocket protein p107 is also preferentially phosphorylated by cdk4/D1, Although Rb and p107 differ significantly in sequence, the Rb Ser795 site can replace the p107 Ser842 site without affecting the rate of phosphorylation, These results suggest that although a determinant of specificity resides in the sequences surrounding the phosphorylated site, the structural context of the site is also a critical parameter of specificity.