人红细胞内血红蛋白与硫氧还蛋白过氧化物酶2的相互作用  

Study on the interaction between hemoglobin and peroxiredoxin 2 using electrophoresis release test

作　　者：解彩霞[1] 安亮[1,2] 魏春华[1] 李晓晶[1] 杜晓红[2] 李嘉欣[1] 苏燕[1] 

机构地区：[1]包头医学院生物化学与分子生物学教研室,内蒙古包头014060 [2]包头医学院第三附属医院检验科

出　　处：《中国输血杂志》2015年第12期1444-1447,共4页Chinese Journal of Blood Transfusion

基　　金：国家自然科学基金(81160214);内蒙古自然科学基金(2010BS1101);内蒙古自治区卫生厅基金(2010056);内蒙古自治区研究生教育创新项目(S201310127(Y02);S201410127(Y01))

摘　　要：目的研究人红细胞内硫氧还蛋白过氧化物酶2(Prx2)与血红蛋白(Hb)的相互作用。方法首先利用红细胞Hb再释放试验分离红细胞和溶血液电泳释放的4个区带,即Hb A、Hb A2、Hb A与Hb A2之间(Hb A-A2)及再释放Hb,将这些区带分别切下,通过反复冻融获得各区带所含蛋白;用葡聚糖G-25浓缩后再利用SDS-PAGE分离各区带所含蛋白组分,并用抗-Prx2做免疫印迹分析;最后用微量热泳动试验(MST)分别检测Prx2蛋白与Hb A和Hb A2之间的结合常数。结果红细胞电泳释放的区带Hb A、Hb A2、Hb A-A2及再释放Hb中均有Prx2;而溶血液电泳释放的Hb区带中,除Hb A2外其余各Hb区带中均有Prx2。MST检测显示,Prx2蛋白与Hb A的结合常数(Kd值)为(14±1.08)μmol/L,而与Hb A2之间的Kd值未被检测到。结论 Prx2与Hb A之间存在相互作用,但与Hb A2之间可能没有相互作用。Objective To study the interaction between peroxiredoxin 2（ Prdx2） and hemoglobin（ Hb） within adult erythrocytes. Methods First,hemoglobin release test（ HRT） was used to separate four Hb bands of the RBC and hemolysate samples. Then,the bands of Hb A,Hb A2,Hb A to Hb A2（ Hb A-A2） and the re-released Hb were cut out from the gel,from which,the proteins were enriched by freeze-thaw method. After further thickening with sephadex-G25,the proteins were separated with 5%- 12% SDS-PAGE and a Western Blot was performed on anti-Prdx2 antibody. Finally,microscale thermophoresis（ MST） was used to test the binding constant between Prdx2,Hb A and Hb A2. Results Prdx2 could be immunoblotted in the Hb A,Hb A2,Hb A-A2 and re-released Hb of the RBC samples,but could only be immunoblotted in the Hb A,Hb A-A2 and re-released Hb of the hemolysate samples. There was no Prdx2 in the Hb A2 of hemolysate sample. MST results showed that the Kd between Hb A and Prdx2 was（ 14 ± 1. 08） μmol / L,but the Kd between Hb A2 and Prdx2 had not been detected. Conclusion There should be interaction between Prdx2 and Hb A,but no interaction between Prdx2 and Hb A2.

关 键 词：红细胞 血红蛋白 HB A HB A2 硫氧还蛋白过氧化物酶2 Hb再释放试验 SDS-PAGE 微量热泳动 

分 类 号：R331.141[医药卫生—人体生理学] R446.112[医药卫生—基础医学]