Microsomal suspensions of liver can catalyze the reduction of cytochrome b5 by the membrane-bound NADPH-cytochrome c reductase but the highly purified NADPH-cytochrome c reductase obtained by lipase solubilization does not reduce cytochrome b5 under normal assay conditions. However, the purified reductase does reduce cytochrome b5 in solutions containing high concentrations of salt. Although the flavoprotein rapidly generates superoxide anion in the presence of high concentrations of salt, the reduction of cytochrome b5 is not dependent upon superoxide anion generation but possibly occurs via a direct interaction between the flavoprotein and the cytochrome. Further, the mechanism of cytochrome b5 reduction involves the cycling of the enzyme between its fully reduced and half-reduced forms during the steady-state reduction of cytochrome. The reduction of cytochrome b5 by detergent- and proteolytically solubilized NADPH-cytochrome c reductase is similar in the presence of high concentrations of KCl. However, with low salt concentrations, only the detergent-solubilized NADPH-cytochrome c reductase reduces cytochrome b5, albeit slowly. No apparent differences in rate of reduction or steady-state level of cytochrome b5 reduction is seen when either the detergent- or proteolytically prepared cytochrome b5 is employed.
ASJC Scopus subject areas
- Molecular Biology