The purification and properties of glutathione peroxidase of erythrocytes.

Previous studies (1,2) have shown the presence of a peroxidase in erythrocytes which catalyzes the breakdown of hydrogen peroxide with reduced glutathione serving as a hydrogen donor. The significance of this enzyme in the protection of hemoglobin from oxidative breakdown has been shown previously (2). The erythrocyte reduced glutathione peroxidase has now been purified by column chromatography on a modified cellulose anion exchange column. The peroxidase activity has been separated completely from the catalase activity, confirming the previous conclusion that the peroxidase and catalase activities of the red cell could be attributed to two distinct enzymes. Although the enzyme in impure preparations is relatively stable, purification results in a markedly increased lability. In addition, some systematic studies have been carried out with this purified enzyme preparation to determine the enzyme specificity in regard to hydrogen donors. Plant peroxidases, in general, have a relatively low degree of specificity since a wide variety of compounds, including organic amines, aromatic diols, and sulfhydryl compounds, will serve as hydrogen donors (3-6). In contrast to this, certain animal peroxidases, such as tryptophan peroxidase, show considerable substrate specificity (4). Since reduced glutathione peroxidase differs from most other perosidase enzymes in that cyanide and azide fail to inhibit its activity (l), it appears likely that there might be differences in specificity for hydrogen donors. In the present st.udy, the enzyme specificity of reduced glutathione peroxidase has been studied from several different aspects. First, we have studied the nature of the hydrogen donor which is necessary for the reduction of hydrogen peroxide in the presence of peroxidase. In addition, other hydrogen donors have been utilized in place of ascorbic acid in the reaction with oxyhemoglobin to yield hydrogen peroxide, and the ability of the peroxidasereduced glutathione system to protect hemoglobin from oxidation under these conditions has been determined.