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OALib Journal期刊
ISSN: 2333-9721
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重组gst-crosstide蛋白的表达纯化及其在agc型蛋白激酶活性检测中的应用
DOI: 10.3724/SP.J.1145.2014.00223, PP. 223-226
Keywords: gst-crosstide,agc型蛋白激酶,体外磷酸化,akt1,ypk1,pkh1
Abstract:
agc型蛋白激酶在信号通路中占据着极为重要的地位，通过对各种信号通路的调控来影响细胞的生长、增殖和凋亡进程.为方便检测agc型蛋白激酶的活性，设计并构建了新型agc型蛋白激酶特异底物gst-crosstide.通过同位素标记体外磷酸化的方法研究了酵母表达的pkh1、ypk1和大肠杆菌表达的akt1三种agc型蛋白激酶磷酸化gst-crosstide的能力.结果显示，酵母表达的有活性的pkh1和ypk1均可有效磷酸化gst-crosstide，并且由于pkh1对ypk1的进一步激活，二者共同作用可显著增加gst-crosstide磷酸化.而仅采用大肠杆菌表达的没有被活化的akt1则无法磷酸化gst-crosstide，但akt1可以被pkh1或ypk1激活，从而获得磷酸化gst-crosstide的能力.研究表明，gst-crosstide可以作为agc蛋白激酶活性检测的有效工具，并将有效促进真核生物蛋白激酶调控的细胞信号转导通路的研究.
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