葡萄糖-6-磷酸脱氢酶缺乏时红细胞膜带3蛋白酪氨酸磷酸化的改变

Changes of protein tyrosine phosphorylation in erythrocyte band 3 glucose-6-phosphate dehydrogenase deficiency

目的 从红细胞膜蛋白磷酸化改变的角度探讨葡萄糖 6 磷酸脱氢酶 (G6PD)缺乏症溶血的机制。方法 Westernblot法检测G6PD缺乏症的红细胞膜蛋白磷酸化的改变以及二硫苏糖醇 (DTT)对蛋白磷酸化的影响 ;以对硝基苯磷酸 (PNPP)为底物 ,测磷酸酪氨酸磷酸酶 (PTPs)活性以探讨磷酸化改变的可能成因。结果 G6PD缺乏的红细胞膜带 3(Band 3)蛋白酪氨酸的磷酸化水平较正常对照明显增多 ,而PTPs活性检测较正常对照明显减弱 ;DTT处理的G6PD缺乏红细胞 ,其膜Band 3蛋白酪氨酸的磷酸化与未处理者无明显差异 ,PTPs活性检测结果与未处理者亦无显著差异。结论 氧化致使G6PD缺乏红细胞的PTPs活性减弱 ,膜Band 3蛋白酪氨酸磷酸化增强 ,成为红细胞溶血的一个重要原因。但PTPs巯基的改变并不是影响PTPs活性的唯一因素。

Objective To explore the hemolytic mechanism of glucose 6 phosphate dehydrogenase(G6PD) deficient erythrocytes in the view of phosphorylation of membrane protein. Methods The alternation of membrane protein phosphorylation and the effect of dithiothreitol(DTT) on protein phosphorylation were analysed by Western blot technique. The activity of phosphotyrosine phosphatase(PTPs) was determined by using p nitrophenyl phosphate as substrate. Results Tyrosine phosphorylation of band 3 protein was obviously enhanced in G6PD deficient erythrocytes. The activity of PTPs was low compared to the normal erythrocytes. The level of phosphotyrosine in G6PD deficient erythrocytes incubated with DTT was almost the same as in those without DTT. The results were consistent with the activity of PTPs. Conclusions PTPs activity reduction and tyrosine phosphorylation enhancement induced by oxidation in G6PD deficiency play an important role in erythrocytes hemolysis. However, the alternation of thiol group is not the only factor affecting the activity of PTPs in G6PD deficient erythrocytes.