Premium
Molecular interactions of re‐released proteins in electrophoresis of human erythrocytes
Author(s) -
Su Yan,
Shen Jing,
Gao Lijun,
Tian Huifang,
Tian Zhihua,
Qin Wenbin
Publication year - 2012
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201100644
Subject(s) - band 3 , agarose , electrophoresis , carbonic anhydrase , chemistry , gel electrophoresis , carbonic anhydrase ii , biochemistry , chromatography , enzyme , membrane protein , membrane
Recently, we found that hemoglobin ( H b) could be re‐released from live erythrocytes during electrophoresis release test ( ERT ). The re‐released H b displays single‐band and multiple‐band re‐release types, but its exact mechanism is not well understood. In this article, the protein components of the single‐band re‐released H b were examined. First, the re‐released band of erythrocytes and the corresponding band of hemolysate, which was used as control, were cut out from starch‐agarose mixed gel. Next, proteins were recovered from the starch‐agarose mixed gel by freeze‐thaw method. After condensing in a vacuum freeze drier, the samples were loaded onto a 5–12% SDS‐PAGE . After electrophoresis, three protein bands (16, 28.9, and 29.3 k D a) emerged from the erythrocytes re‐released H b single‐band ( R ‐ R ), but only one band (29.3 k D a) emerged from the corresponding hemolysate control band ( H ‐ R ). Finally, these bands were analyzed by MALDI – TOF MS . The results showed that these proteins were beta‐globin (16 k D a), carbonic anhydrase 1 ( CA 1, 28.9 k D a), and carbonic anhydrase 2 ( CA 2, 29.3 k D a). Because CA 2 exists in both erythrocytes re‐released band and hemolysate control band, we conclude that the single‐band re‐released H b is mainly composed of H b A and CA 1. Studying the possible interaction between H b A and CA 1 will help us further understand the in vivo function of H b.