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目的:研究人肝癌细胞株SMMC-7721中转醛醇酶(Transaldolase,TAL)活性变化与磷酸化的关系。方法:SMMC-7721细胞培养,细胞周期同步化后分组加不同浓度(10%、1%)胎牛血清孵育,分别测定不同孵育时间(1、2、4、8h)TAL酶活性;Western blot检测TAL蛋白表达水平。磷酸酶消化法检测TAL活性变化与磷酸化的关系。结果:①人肝癌细胞株SMMC-7721中TAL活性在10%胎牛血清培养时明显高于1%胎牛血清,2、4、8h显著性差异(P<0.05)。②不同浓度(10%.1%)胎牛血清孵育2、4、8hTAL蛋白表达水平无显著性差异。③磷酸化消化后TAL活性,与消化前相比呈显著性下降。结论:人肝癌细胞株SMMC-7721中增加血清浓度明显提高TAL活性,酶活性变化与TAL蛋白表达水平无关,提示活性调节发生在蛋白翻译后修饰,其中磷酸化起重要作用。
Objective: To study the relationship between the changes of phosphorylation and the activity of transaldolase (TAL) in human hepatocellular carcinoma cell line SMMC-7721. Methods: SMMC-7721 cells were cultured and synchronized with different cell cycle (10%, 1% fetal bovine serum). TAL activity was measured at different incubation times (1, 2, 4, 8h) TAL protein expression level. Relationship between TAL activity and phosphorylation by phosphatase digestion. Results: ① The TAL activity of human hepatocellular carcinoma cell line SMMC-7721 was significantly higher than that of 1% fetal bovine serum in 10% fetal bovine serum (P <0.05). ② There was no significant difference in the expression of TAL protein at 2,4,8 and 8 h after incubation with different concentrations of fetal bovine serum (10%, 1%). ③ phosphorylation TAL activity after digestion, compared with before digestion was significantly decreased. CONCLUSION: Increased serum concentration of human hepatocellular carcinoma cell line SMMC-7721 can significantly increase the activity of TAL. The change of enzyme activity has nothing to do with the expression of TAL protein, suggesting that the regulation of the activity occurs in the post-translational modification of protein. Phosphorylation plays an important role.