Stoichiometry of O2 Metabolism and NADPH Oxidation of the Cell-free Latent Oxidase Reconstituted from Cytosol and Solubilized Membrane from Resting Human Neutrophils*

摘要: The stoichiometry of NADPH-dependent O2 consumption expressed by reconstituted latent oxidase obtained combining cytosol and membrane fractions from resting human neutrophils with GTP gamma S SDS in a cell-free assay was evaluated regard to NADPH consumed superoxide H2O2 production. Oxidase activity monitored simultaneously uptake analysis using Clark cell electrode for consumption, spectrally at 340 nm oxidation, the presence excess dismutase catalase, yielded an uptake: ratio 0.51 +/- 0.04 (+/- 1 S.D., n = 6). In varying concentrations ferricytochrome c 100 microM, exclusion dismutase, net rate plateaued approximately 6% seen final mixtures. Cytosol solubilized employed these assays were devoid endogenous trapping, or dismutase-like, activities. These results indicate that total consumed, 94% is associated direct univalent generation superoxide. remaining albeit low level appears be recovered form indicating capable channeling electron equivalents through its redox sites highly controlled manner ensuring principal reduction product concomitant oxidation NADPH.