SELENITE IS A SUBSTRATE FOR CALF THYMUS THIOREDOXIN REDUCTASE AND THIOREDOXIN AND ELICITS A LARGE NONSTOICHIOMETRIC OXIDATION OF NADPH IN THE PRESENCE OF OXYGEN

The thioredoxin system, comprising NADPH, thioredoxin reductase and thioredoxin reduces protein disulfides via redox-active dithiols. We have discovered that sodium selenite is a substrate for the thioredoxin system; 10-mu-M selenite plus 0.05-mu-M calf thymus thioredoxin reductase at pH 7.5 caused a non-stoichiometric oxidation of NADPH (100-mu-M after 30 min). In contrast, thioredoxin reductase from Escherichia coli showed no direct reaction with selenite, but addition of 3-mu-M E. coli thioredoxin also resulted in non-stoichiometric oxidation of NADPH, consistent with oxidation of the two active-site thiol groups in thioredoxin to a disulfide. Kinetically, the reaction was complex with a lag phase at low selenite concentrations. Under anaerobic conditions the reaction stopped after 1 mol selenite had oxidized 3 mol NADPH; the admission of air then resulted in continued consumption of NADPH consistent with autooxidation of selenium intermediate(s). Ferricytochrome c was effectively reduced by calf thymus thioredoxin reductase and selenite in the presence of oxygen. Selenite caused a strong dose-dependent inhibition of the formation of thiol groups from insulin disulfides with either the E. coli or calf-thymus thioredoxin system. Thus, under aerobic conditions selenite catalyzed, NADPH-dependent redox cycling with oxygen, a large oxygen-dependent consumption of NADPH and oxidation of reduced thioredoxin inhibiting its disulfide-reductase activity.