Regulation of Pyruvate Dehydrogenase in Muscle

Abstract

Abstract The active and inactive forms of pyruvate dehydrogenase were assayed in a mitochondrial extract prepared from rat skeletal muscle. The active form of this enzyme was increased almost 3-fold upon preincubation with 10 mm Mg++ and Ca++. There was a decrease of approximately 30% when 1 mm ATP and 0.5 mm Mg++ were present in the original preincubation mixture. If citrate (5.0 mm) was added to the preincubation mixture containing 10 mm Mg++ and Ca++, the activation of pyruvate dehydrogenase was almost completely prevented. The concentration of citrate required to reduce the rate of activation of pyruvate dehydrogenase by 50% was approximately 2.0 mm in this system. Citrate did not inhibit the assay system for activated pyruvate dehydrogenase. Once the activation of pyruvate dehydrogenase was complete, addition of 5 mm citrate to the preincubation mixture did not decrease this activity. Increasing the concentrations of Mg++ and Ca++ from 10 to 20 mm did not overcome the citrate (5 mm)-induced prevention of pyruvate dehydrogenase activation. Only the citrate analogues with a hydroxy group (citrate, (-) hydroxycitrate, fluorocitrate, 2-ethylcitrate and 2-methylcitrate) inhibited the activation of pyruvate dehydrogenase independent of Mg++ and Ca++ concentration, whereas this was not the case with tricarboxylic acids without a hydroxy group (tricarballylate and 1,2,3-tricarboxybenzene). It is concluded that citrate can prevent the activation of pyruvate dehydrogenase by a mechanism independent of Mg++ and Ca++ chelation. Some possible physiological implications are discussed.