Interactions between peroxiredoxin 2, hemichrome and the erythrocyte membrane

Abstract

Peroxiredoxin 2 (Prx2) is an abundant antioxidant protein in erythrocytes that protects against hemolytic anemia resulting from hemoglobin oxidation and Heinz body formation. A small fraction of Prx2 is bound to the cell membrane, but the mechanism and relevance of binding are not clear. We have investigated Prx2 interactions with the erythrocyte membrane and oxidized hemoglobin and whether these interactions are dependent on Prx2 redox state. Membrane binding of Prx2 in erythrocytes decreased when the cells were treated with HO, but studies with purified Prx2 and isolated ghosts showed that the interaction was independent of Prx2 redox state. Hemoglobin oxidation leads to the formation of hemichrome, a denatured form of the protein that binds to Band3 protein in the cell membrane as part of the senescence process and is a precursor of Heinz bodies. Hemichrome competed with Prx2 and decreased Prx2 binding to the membrane, potentially explaining the decreased binding in oxidant-exposed cells. The increased membrane binding of Prx2 seen with increasing intracellular calcium was less sensitive to HO or hemichrome, suggesting an alternative mode of binding. Prx2 was also shown to exhibit chaperone-like activity by retarding the precipitation of pre-formed hemichrome. Our results suggest that Prx2, by restricting membrane binding of hemichrome, could impede Band3 clustering and exposure of senescence antigens. This mechanism, plus the observed chaperone activity for oxidized hemoglobin, may help protect against hemolytic anemia.

The Effect of Sepsis on the Erythrocyte

“…A second RBC peroxidase is the thiol-dependent peroxiredoxin II [ 104 , 105 , 106 ] (Prx2 or PRDX2), the third most abundant protein in the RBC, of which a small fraction binds to the inner leaflet of the cell membrane [ 105 ]. The high concentration of Prx2 in the RBC suggests that it functions to detoxify low levels of H 2 O 2 that are generated through autoxidation mechanisms [ 107 ].…”

Interplay between Erythrocyte Peroxidases and Membrane

“…Thus, they demonstrated that Prx2 prevents metHb aggregation, and, probably acts as a chaperone for the denatured Hb [96]. Contrary to what was previously observed [26,31,32], Bayer et al [96] found a decrease in Prx2 membrane binding, with increasing concentrations of H 2 O 2 . A decrease in Prx2 linkage to the RBC membrane with OS conditions was also observed in beta-thalassemic mice RBCs, probably due to the increase in metHb that binds to the membrane, reducing the access of Prx2 to the same site [99].…”

“…CAT, GPx and Prx2 are essentially cytosolic enzymes; however, the association of these enzymes to the erythrocyte membrane has been reported in different in vivo and in vitro studies [26,31,32,[93][94][95][96][97]. Erythrocytes from patients with hereditary spherocytosis showed CAT [95] and Prx2 bound to their membranes [26].…”

“…In recent in vitro assays performed by our group [32], we showed that H 2 O 2 -induced oxidative stress triggered the binding of Prx2 and GPx to RBC membrane. A recent study by Bayer et al [96] about the interaction of Prx2 with the RBC membrane reported that the linkage of Prx2 to the membrane is independent of its redox state and that Prx2 competes with Hb for the same binding site in the RBC membrane. Thus, they demonstrated that Prx2 prevents metHb aggregation, and, probably acts as a chaperone for the denatured Hb [96].…”

“…A recent study by Bayer et al [96] about the interaction of Prx2 with the RBC membrane reported that the linkage of Prx2 to the membrane is independent of its redox state and that Prx2 competes with Hb for the same binding site in the RBC membrane. Thus, they demonstrated that Prx2 prevents metHb aggregation, and, probably acts as a chaperone for the denatured Hb [96]. Contrary to what was previously observed [26,31,32], Bayer et al [96] found a decrease in Prx2 membrane binding, with increasing concentrations of H 2 O 2 .…”