Partition of catalase and its peroxidase activities in human red cell membrane: effect of ATP depletion.
Partititon of catalase (hydrogen-peroxide:hydrogen-peroxide oxidoreductase EC 1.11.1.6) and peroxidase (donor:hydrogen-peroxide oxidoreductase EC 1.11.1.7) activities between the red cell membrane and the cytosol were studied under various experimental conditions. A small but significant amount of catalase (1.6%) was retained on human red cell membranes prepared by hemolysing washed red cells with 30 volumes of 10 mM Tris buffer, pH 7.4. Membrane -bound catalase had a relatively higher peroxidase activity than the soluble enzyme fraction. Polyacrylamide gel electrophoresis in sodium dodecyl sulfate of the solubilized membranes demonstrated catalase to be a single band with a molecular weight of 60 000. Membranes prepared from adenosine triphosphate-depleted red cells depicted a two to three-fold increase in catalase activity, as well as an increase in 60 000 molecular weight band on polyacrylamide gel electrophoresis. The extra amount of retained catalase was a less efficient peroxidase than found in fresh membranes. The binding of catalase to ATP-depleted red cell membranes was dependent upon both pH and hemolysing ratio. Red cells incubated at pH 7.1 demonstrated a decrease in bound catalase, as did membranes prepared from red cells hemolysed at 1:100 dilution. beta-Mercaptoethanol decreased the catalase activity in the membranes and increased the odianisidine peroxidase activity without any significant effect on the 60 000-dalton band.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism*; D004295:Dihydroxyphenylalanine / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004910:Erythrocyte Membrane / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006461:Hemolysis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008623:Mercaptoethanol / Q000494:pharmacology; D008970:Molecular Weight; D010544:Peroxidases / Q000378:metabolism*
D008623:Mercaptoethanol; D004295:Dihydroxyphenylalanine; D000255:Adenosine Triphosphate; D010544:Peroxidases; D002374:Catalase