Role of Enzyme-Enzyme Interactions in the Regulation of Gluconeogenesis

Mitochondria isolated from rat kidney inactivated the soluble gluconeogenic enzyme, d-fructose 1,6-diphosphatase, when they were incubated with the purified enzyme in the presence of ATP, magnesium, and cysteine. The addition of malate, α-ketoglutarate, pyruvate, succinate, myristate, palmitate, stearate, arachidate, or oleate to the reaction mixture decreased the rate of inactivation of d-fructose 1,6-diphosphatase, whereas the addition of 2,4-dinitrophenol or pentachlorophenol increased the amount of enzyme inactivated. The highest rates of inactivation were obtained when ADP was added in place of ATP. The rate of conversion of the enzyme to an inactive form increased with increasing concentrations of adenine nucleotide and mitochondria. The oxidation of tricarboxylic acid cycle intermediates and fatty acids increases the ratio of ATP to ADP inside the mitochondria, and this high ratio appears to inhibit the activity of the regulatory system. The 2,4-dinitrophenol, on the other hand, lowers the ratio of ATP to ADP in the mitochondria by uncoupling oxidative phosphorylation and thereby increases the activity of the inactivating system. The administration of fatty acids and tricarboxylic acid cycle intermediates stimulate gluconeogenesis in vivo, and the results of the present study indicate that this effect may be due, in part, to the inhibition of a regulatory enzyme system present in mitochondria which inactivates d-fructose 1,6-diphosphatase when an oxidizable substrate is present to maintain a high ratio of ATP to ADP in the mitochondria.