Mechanisms of Adriamycin-Dependent Oxygen Activation Catalyzed by NADPH-Cytochrome c-(Ferredoxin)-Oxidoreductase

Abstract

Under aerobic conditions, O2 uptake and production of O2 - and H2O2 by isolated NADPH - cytochrome c-(ferredoxin)-oxidoreductase from Euglena gracilis was strongly stimulated by adriamycin. Further stimulation was not observed with 0.1 mᴍ Fe3+-EDTA. Methionine fragmentation (measured as ethylene release), as a reliable indicator for the formation of OH- radical-like oxidants under aerobic conditions (100 μmol O2 in a 10 ml reaction vessel) was strongly stimulated by 0.1 mᴍ Fe3+-EDTA or, in the absence of iron, by partial anaerobiosis (1 μmol O2 per vessel). The highest rate of methionine fragmentation was observed under anaerobic conditions in the presence of both reduced adriamycin and added H2O2. Aerobic methionine fragmentation in the presence of adriamycin and Fe3+-EDTA was inhibited by superoxide dismutase and catalase by more than 90%, while methionine fragmentation under semianaerobiosis in the absence of Fe3+-EDTA was inhibited by superoxide dismutase to only about 50%, while catalase again inhibited by more than 90%. These results indicate that the adriamycin-catalyzed production of a strong oxidant appears to be governed by different mechanisms depending on oxygen availability; namely the production of a Fenton-type oxidant driven by adriamycin-catalyzed superoxide formation and also, the formation of the “crypto-OH- radical” by direct electron donation from the adriamycin semiquinone radical to H2O2 under oxygen limiting conditions