【摘要】 目的 :利用PKCε酶促放射性磷酸化标记兔心肌细胞蛋白质 ,鉴定出其可能的直接底物 ,为进一步研究奠定基础 .结果 :通过PKCε特异底物竞争抑制、PKCε特异抑制剂Ro 31- 82 2 0、兔心肌细胞蛋白质热变性等分析措施 ,初步用PKCε酶促放射性磷酸化标记 ,从兔心肌细胞蛋白质中鉴定出表观分子量为 5 7kd ,35kd ,2 3kd和 2 0kd的蛋白质的磷酸化与PKCε酶活性相关 ,其中 5 7kd ,2 3kd和 2 0kd 3种蛋白质可被PKCε在体外直接磷酸化 ,并且其磷酸化不受蛋白激酶抑制剂PD980 5 9和SB2 0 35 8的影响 .结论 :5 7kd ,2 3kd和2 0kd 3种蛋白质有可能是PKCε直接底物 .更多还原

【Abstract】 Ischemic preconditioning has protective effect on ischemic heart. Protein kinase Cε(PKCε) plays a key role in the intracellular signal transduction pathway of such protective mechanism. But the direct downstream substrate of PKCε related to ischemia preconditioning stays to be identified. In this study, the phosphorylated proteins by PKCε in rabbit cardiomyocyte lysate were examined. At least 4 of phosphorylated proteins with appear molecule weights of 57?kd, 35?kd, 23?kd, and 20?kd are related to PKCεs kinase activity through inhibition assays of PKCε special substrate and PKC special inhibitor Ro 31-8220. The protein kinase inhibitors(PD98059 and SB20358) did not show any inhibitive effects on the phosphorylation of these proteins. Among them, the 57?kd, 23?kd, and 20?kd proteins are directly phosphorylated when the kinase activity in the lysate was eliminated by heating denaturation and the phosphorylation assay was carried out with adding of purified recombinant PKCε.They are potential direct substrates of PKCε in rabbit cardiomyocyte.更多还原