Two Forms of Adrenodoxin Reductase from Mitochondria of Bovine Adrenal Cortex

Two forms of adrenodoxin reductase, differing in isoelectric properties and thermal stability, have been highly purified from bovine adrenal cortex mitochondria. Both preparations appear to be homogeneous based upon polyacrylamide disc‐gel electrophoresis, isoelectric focusing, and ion‐exchange chromatography. Both are monomeric proteins containing one FAD per molecule (Mr 50000) and exhibit spectra typical of flavoproteins. The two preparations are indistinguishable by their amino acid compositions and N‐terminal (serine) and C‐ter‐ minal (leucine) amino acids, but reveal distinct sugar contents. Electrophoretic mapping of the protease digests of the two preparations indicates that the two have identical, or closely similar, primary structures. Each form functions equally as an adrenodoxin‐dependent electron transport component from NADPH to the P‐450 cytochromes in the two different reconstituted steroid monooxygenase systems which catalyze cholesterol side‐chain cleavage reaction and steroid 11β‐(18‐)hydroxylase reaction, respectively. In all adrenodoxin reductase‐ containing fractions from adrenal cortex so far examined, i.e. the soluble fraction, and the sonic‐solubilized and detergent‐solubilized mitochondrial fractions, a constant ratio of about 2:5 between the two forms is found as judged by the DEAE‐cellulose chromatogram. No evidence has been obtained for mutual conversion of the two forms of the reductase, or for dissociation of the sugar moiety during the purification process and under various conditions examined.