Microsomal Electron Transport

NADPH-cytochrome c reductase in rat liver microsomes was solubilized by bromelain digestion and purified to homogeneity. An antibody preparation obtained by immunization with this enzyme was found to inhibit the NADPHcytochrome c reductase activity of both the purified enzyme and intact microsomes by more than 90%. This antibody also inhibited the NADPH-dependent peroxidation of microsomal lipid which occurs in the presence of ferric ion chelated by ADP (ADP-Fe). The bromelain-solubilized reductase will also promote NADPH-dependent peroxidation of extracted microsomal lipid, but only if ferric ion chelated by ethylenediaminetetraaacetate (EDTA-Fe) is included in the reaction mixture. The optimal conditions for this reaction are those required for the optimal reduction of EDTA-Fe by the purified reductase. Other ferric ion chelators, regardless of whether or not they promoted the reduction of ferric ion, would not replace EDTA in promoting lipid peroxidation. A purified preparation of the microsomal enzyme, NADHcytochrome bS reductase, which reduces EDTA-Fe, will also promote the peroxidation of extracted microsomal lipid. Intact microsomes, in the presence of ADP-Fe, are specific for NADPH instead of NADH in promoting the peroxidation of microsomal lipids; however, in the presence of both EDTA-Fe and ADP-Fe, both NADH and NADPH promote lipid peroxidation. These results indicate that the NADPHdependent peroxidation of microsomal lipid involves the activity of NADPH-cytochrome c reductase, and suggest that an additional microsomal electron transport component is involved. The function of this additional component in the lipid peroxidation reaction can apparently be replaced by EDTA-Fe.