POSSIBLE TARGET COMPONENTS FOR THE INHIBITORY EFFECT OF N-ETHYLMALEIMIDE ON THE ACTIVATION OF NEUTROPHIL NADPH OXIDASE

Potential target components for the inhibitory effect of covalent sulf hydryl-modifying reagent N-ethylmaleimide (NEM) on the activation of N ADPH oxidase in human neutrophils was studied in a cell-free system. T he capacity of both cytosol and membrane fractions to induce the trans location of cytosolic components and O-2(-) generation in the cell-fre e activation system was affected by NEM. The phosphorylation of p47(ph ox), which mediates the translocation of cytosolic complex, by protein kinase C was not inhibited by NEM and NEM-treated p47(phox) was as ef fective as untreated p47(phox) both in the kinase-dependent and in the amphiphile-dependent cell-free activation sysyems. In addition, phosp hatidic acid-dependent phosphorylation of cytosol including p47(phox) was not affected by NEM. The inhibition of cytosol's capacity to activ ate NADPH oxidase was partially reversed by an addition of the fractio n containing G-protein vac. Taken together, the data suggest that memb rane component cytochrome b(558) and cytosolic component vac may be th e potential targets for the NEM effect on the activation of NADPH oxid ase.