A Function of Cytochrome b5 in Fatty Acid Desaturation by Rat Liver Microsomes

Abstract

On addition of NADPH to rat liver microsomes cytochrome b₅ assumes a steady-state reduction level, based on a balance between its reduction by NADPH and its aut-oxidation. Initiation of fatty acid desaturation by adding stearoyl-CoA (stearyl CoA) results in a rapid shift of the NADPH-supported steady state of the cytochrome in favor cf more oxidation, and this shift is intensified by partially inhibiting microsomal NADPH-specific flavoprotein by H_gCl₂. This suggests that increased utilization of reducing equivalents for desaturation is accompanied by a stimulation of reoxidation of cytochrome bt. Actually, the magnitude of stearyl CoA-induced shift is roughly proportional to the fatty acid desaturation activity of the microsomes employed. Moreover, this shift is interfered with by cyanide which inhibits a terminal component (“cyanide-sensitive factor”) of the desaturation system. Since the reduction of microsomal bound cytochrome bt by NADH is much faster than the autoxidation of the cytochrome and the overall desaturation reaction, NADH causes complete reduction of cytochrome b₅ and no change in the reduction level is observed on addition of stearyl CoA. However, when NADH-cytochrome b₅ reductase is strongly inhibited by PCMS, stearyl CoA does lower the steady-state reduction level of cytochrome b₅ in NADH-treated microsomes. Stearyl CoA also stimulates the oxidation of cytochrome b₅ observable on exhaustion of the NADH added, and this stimulation is again prevented by cyanide. With microsomes from which cytochrome b₅ has been removed to various extents by mild proteolytic digestion, the desaturation of stearyl CoA supported by ascorbate as electron donor is dependent on the content of remaining cytochrome b₅. It is concluded that cytochrome b₅ in liver microsomes acts as an intermediary electron carrier which passes reducing equivalents from NADH, NADPH and ascorbate to the cyanide-sensitive factor where fatty acid desaturation per se probably takes place. Microsomes prepared from rat epididymal adipose tissue contain a significant amount of cytochrome b₅ which plays a similar role in fatty acid desaturation.