Mitochondrial respiration and membrane potential are regulated by the allosteric ATP-inhibition of cytochrome c oxidase

This paper describes the problems of measuring the allosteric ATP-inhibition of cytochrome c oxidase (CcO) in isolated mitochondria. Only by using the ATP-regenerating system phosphoenolpyruvate and pyruvate kinase full ATP-inhibition of CcO could be demonstrated by kinetic measurements. The mechanism was proposed to keep the mitochondrial membrane potential (∆Ψm) in living cells and tissues at low values (100–140 mV), when the matrix ATP/ADP ratios are high. In contrast, high ∆Ψm values (180–220 mV) are generally measured in isolated mitochondria. By using a tetraphenyl phosphonium electrode we observed in isolated rat liver mitochondria with glutamate plus malate as substrates a reversible decrease of ∆Ψm from 233 to 123 mV after addition of phosphoenolpyruvate and pyruvate kinase. The decrease of ∆Ψm is explained by reversal of the gluconeogenetic enzymes pyruvate carboxylase and phosphoenolpyruvate carboxykinase yielding ATP and GTP, thus increasing the matrix ATP/ADP ratio. With rat heart mitochondria, which lack these enzymes, no decrease of ∆Ψm was found. From the data we conclude that high matrix ATP/ADP ratios keep ∆Ψm at low values by the allosteric ATP-inhibition of CcO, thus preventing the generation of reactive oxygen species which could generate degenerative diseases. It is proposed that respiration in living eukaryotic organisms is normally controlled by the ∆Ψm-independent “allosteric ATP-inhibition of CcO.” Only when the allosteric ATP-inhibition is switched off under stress, respiration is regulated by “respiratory control,” based on ∆Ψm according to the Mitchell Theory.

Research Highlights
► Regulation of mitochondrial respiration.
► Regulation of mitochondrial membrane potential.
► Regulation of Cytochrome c oxidase activity.
► Phosphorylation of Cytochrome c oxidase subunits.