The effects of disruption of genes for peroxiredoxin-2, glutathione peroxidase-1, and catalase on erythrocyte oxidative metabolism

Since 1-2% of the cell's oxyhemoglobin undergoes spontaneous heterolytic dissociation into metHb and superoxide every day [1], and superoxide is readily converted to H2O2 by superoxide dismutase, the red cell is unavoidably exposed to these reactive oxidizing species (ROS). The survival of the erythrocyte therefore depends on an efficient defense against oxidative damage by H2O2 and superoxide. Three enzymes are believed to participate in the oxidative defense of the red cell: glutathione peroxidase-1 (Gpx1), catalase, and peroxiredoxin-2 (Prdx2). We have begun to examine the function of each of these enzymes using a series of mice in which the gene for one or more of these enzymes has been disrupted. Experiments [2] with the catalase inhibitor 3-amino-1,2,4-triazole (3-AT) found that very little endogenously produced H2O2 is catabolized by catalase in wild-type mouse red cells, but that deletion of Gpx1 increased H2O2 flux through catalase, indicating that Gpx1 plays a major role in eliminating endogenous H2O2. In addition, we were able to show that Gpx1 protects the erythrocyte against attack by organic peroxides [3], as proposed many years ago by Flohe [4]. These experiments with Gpx1 deficient cells, together with a detailed kinetic model of the cell's oxidative metabolism [2], found that the activity of catalase and glutathione peroxidase are inadequate to explain the oxidative catabolism of the red cell, and that peroxiredoxin activity had to be included in the model to explain the experimental results. We here report experiments testing this proposal. The major peroxiredoxin of the mature red cell is Prdx2 [5]. Mice with a disrupted Prdx2 gene have been constructed [6]. These animals are anemic and show various red cell abnormalities. We found that deletion of Prdx2 led to an increase in erythrocyte H2O2, confirming a role for Prdx2 in the cell's endogenous H2O2 metabolism. The loss of Prdx2 had no effect on the ability of the mature red cell to detoxify exogenous H2O2. In addition, although Prdx2 is able to reduce organic peroxides in vitro, it appears not to do this in the red cell. Rather, Gpx1 has the primary role in detoxifying organic peroxides. The data also confirm that exogenous H2O2 is detoxified primarily by catalase. The presence of high levels of Prdx2 together with the high rate of reaction between Prdx2 and H2O2 has led to proposals that Prdx2 is a major antioxidant protein in the red cell [6,7]. Our data suggest that erythrocytic Prdx2 participates in the disposal of endogenous H2O2, but plays no role in eliminating exogenous peroxides. This relatively minor role for Prdx2 in the mature red cell, despite its high concentration, lends support to the proposal [5] that it has its greatest importance in hematopoesis.