蛋白激酶的相互作用促进τ的阿尔茨海默样磷酸化

Interaction of Protein Kinases Stimulates an Alzheimer like Phosphorylation of τ

cＡＭＰ依赖性蛋白激酶（ＰＫＡ） 的预处理可显著增强糖原合成酶激酶３ （ＧＳＫ３） 对τ蛋白的磷酸化作用．将磷酸化的τ蛋白经胰蛋白酶消化， ＦｅＣｌ３ 亲和柱分离及Ｃ１８ 反相高压液相层析纯化后， 再用高压电泳， 手工Ｅｄｍａｎ 降解及自动氨基酸序列分析等技术， 对其磷酸化位点进行鉴定． 结果发现： ＧＳＫ３ 可使ＰＫＡ 预处理的τ至少在丝氨酸（Ｓｅｒ） １９５ ， Ｓｅｒ１９８ ， Ｓｅｒ１９９ ， Ｓｅｒ２０２ ， Ｓｅｒ２３５ ， Ｓｅｒ２６２ ， Ｓｅｒ３５６ ， Ｓｅｒ４０４ ， 苏氨酸（ Ｔｈｒ） ２０５ 和Ｔｈｒ２３１ 等１０ 个位点发生磷酸化． 其中Ｓｅｒ１９８ ， Ｓｅｒ１９９ ， Ｓｅｒ２０２ ， Ｓｅｒ２３５ ， Ｓｅｒ２６２ ， Ｓｅｒ４０４ ， Ｔｈｒ２０５和Ｔｈｒ２３１ 为Ａｌｚｈｅｉｍｅｒ 病（ＡＤ） τ蛋白的异常磷酸化位点． 上述磷酸化作用高度抑制τ的生物学活性， 提示：ＡＤτ的生物学功能的抑制与Ｓｅｒ１９８ ， Ｓｅｒ１９９ ， Ｓｅｒ２０２ ， Ｓｅｒ２３５ ， Ｓｅｒ２６２ ， Ｔｈｒ２０５ 和Ｔｈｒ２３１ 的磷酸化密切相关， ＰＫＡ 和ＧＳＫ３ 的相互作用可能在ＡＤ

Prephosphorylation of τ by cAMP dependent protein kinase (PKA) significantly stimulated its consecutive phosphorylation catalyzed by glycogen synthetase kinase 3 (GSK 3). After digestion of phosphorylated τ with trypsin, 32 P labeled τ peptides were purified by a sequential FeCl 3 micro affinity column and a C 18 reverse phase high performance liquid chromatography. The results from a combined techniques of high voltage electrophoresis , manual Edman degration and auto gas phase amino acid sequence analysis demonstrated that the phosphorylation sites of τ (pretreated with PKA) by GSK 3 were Ser(serine) 195,Ser 198,Ser 199,Ser 202,Ser 235,Ser 262,Ser 356,Ser 404, Thr(threonine) 205 and Thr 231. Among them, Ser 198,Ser 199,Ser 202,Ser 235,Ser 262,Ser 404,Thr 205 and Thr 231 are abnormal phosphorylation sites of τ found in Alzheimer disease. As the above phosphorylation potently inhibited the biological activity of τ, it was concluded that the phosphorylation of τ at above mentioned Alzheimer sites is critical to the inhibition of its biofunction, and that the interaction of PKA and GSK 3 might be a potential system responsible for the neurofibrillary degeneration seen in Alzheimer disease .