A. Perrin et al., METABOLISM OF MALATE IN BOVINE ADRENOCORTICAL MITOCHONDRIA STUDIED BYC-13-NMR SPECTROSCOPY, European journal of biochemistry, 223(1), 1994, pp. 51-59
C-13-NMR spectroscopy was used to study the metabolism of [C-13]malate
in bovine coupled adrenocortical mitochondria. The most apparent diff
erence between the mitochondria from steroidogenic tissues and mitocho
ndria from other tissues is the presence, in addition to the normal re
spiratory chain, of a second electron-transport system responsible for
steroid hydroxylation. [C-13]malate was synthesized from [C-13]succin
ate by isolated adrenocortical mitochondria. The basic functional susp
ension consisted of oxygenated mitochondria to which were added ADP, i
norganic phosphate (P-i) and [C-13]malate, both in the absence or pres
ence of the steroid substrate, deoxycorticosterone. These mitochondria
synthesized [C-13]citrate and [C-13]pyruvate from [C-13]malate. The C
-13 labeling of these two metabolites demonstrated an important role o
f the malic enzyme and the kinetics depended on the presence of the st
eroid substrate; the citric acid cycle was stopped during the hydroxyl
ation pathway. The addition of cyanide, a strong inhibitor of the resp
iratory chain, confirmed an increased malic enzyme activity when hydro
xylation occurred, since pyruvate was trapped by formation of a cyanoh
ydrin. The relative enzymic activities of malic enzyme and isocitrate
dehydrogenase were compared, both in the absence or presence of the st
eroid substrate, by supplementing the basic suspension with unlabeled
exogenous metabolites, such as pyruvate or oxaloacetate.