Quantitative relationship between adrenal cytochrome P-450 and its reducing system, and their organ specificities in relation to corticoidogenesis

Abstract

After 10 kcycles sonication of the adrenal mitochondrial fraction (800–6000 × g precipitate) of rat, the supernatant fluid at 105000 × g for 30 min was resolved into the cytochrome P-450 fraction as the precipitate, and its reducing system (non-heme iron protein + flavoprotein) as the supernatant fluid at 135000 × g for 120 min. Only when the above two fractions were recombined, 11β- and 18-hydroxylations of deoxycorticosterone occurred in the presence of NADPH, though these were individually inactive. Then, the quantitative relationship between the above two fractions in respect to hydroxylation of deoxycorticosterone and the influence of NADPH supply on the activity of the recombined system were examined. By the same procedure as that applied for the above stated subfractionation of adrenal mitochondria, the two submitochondrial fractions were prepared from the testicular and hepatic mitochondria, respectively, of the same animals. When the two individual fractions of the three glands, respectively, were combined and subjected to the enzyme assays, 11β- and 18-hydroxylations of deoxycorticosterone were demonstrable specifically by recombination of the adrenal P-450 fraction and its reducing system of adrenal origin, but were not associated with the corresponding fractions which originated in either testis or liver.