11β-Hydroxylation and carnitine-dependent fatty acid oxidation in adrenal mitochondria

Abstract

14C-palmitate can be oxidized to 14C-CO 2 in rat adrenal homogenates. Carnitine stimulates this conversion threefold. In the presence of carnitine, ketone bodies are also formed, whereas in its absence, none are detected. Carnitine-palmityl transferase activity can be demonstrated in intact as well as detergent-treated mitochondria. 11β-hydroxylation in isolated rat adrenal mitochondria can be supported by palmitate, ATP and coenzyme A, or palmityl coenzyme A if carnitine is present. The activity supported by palmityl CoA + carnitine or palmitylcarnitine alone is 5–30% of the activity with 10 m m 2-oxoglutarate or 5 m m isocitrate, substrates which produce maximal rates of 11β-hydroxylation. The addition of catalytic amounts of malate (0.2 m m, a level which does not support 11β-hydroxylation) increases the activity with palmitylcarnitine or palmityl CoA + carnitine to levels comparable to those obtained with 2-oxoglutarate and isocitrate. Although fluorocitrate inhibits citrate-supported 11β-hydroxylation, it has no effect on 11β-hydroxylation supported by palmitylcarnitine or palmityl CoA + carnitine in the presence of malate. In contrast, amytal and dinitrophenol (DNP) are markedly inhibitory. Oxidation of long-chain fatty acids in rat adrenal mitochondria occurs predominately via a carnitine-stimulated pathway. This activity is capable of supporting 11β-hydroxylation via a pathway in which the NADH formed during β-oxidation is coupled to the energy-dependent transhydrogenase for the generation of NADPH. Maximal rate of 11β-hydroxylation in isolated mitochondria can be obtained with physiological levels of intermediates in fatty acid oxidation in contrast to the high concentration of citric acid cycle intermediates required.