Influence of the mitochondrial outer membrane and the binding of creatine kinase to the mitochondrial inner membrane on the compartmentation of adenine nucleotides in the intermembrane space of rat heart mitochondria

Abstract

The influence of the mitochondrial outer membrane and that of the binding of creatine kinase to the mitochondrial inner membrane on the compartmentation of adenine nucleotides in the intermembrane space of rat heart mitochondria were investigated under conditions of maximal rates of mitochondrial creatine kinase. To this end, experiments were performed in reconstituted sytems consisting of functionally intact rat heart mitochondria or mitoplasts and pyruvate kinase, both competing at ADP formed by mitochondrial creatine kinase in the presence of high concentrations of adenine nucleotides. Results showed that removal of the mitochondrial outer membrane diminished the compartmentation effects, supporting the relevance of the mitochondrial outer membrane to compartmentation effects in the intermembrane space. Compartmentation effects were clearly seen even in KCl-containing media which release the creatine kinase from the inner membrane, indicating that the localization of the ADP-regenerating enzyme in the mitochondrial intermembrane space within the outer membrane is a prerequisite for dynamic compartmentation of adenine nucleotides. Under these conditions, the binding of creatine kinase to the inner membrane is of minor importance. Lowering the ATP concentrations diminished the extent of AdN compartmentation due to decreased creatine kinase rates. Under these conditions of low AdN fluxes, the release of creatine kinase from the inner membrane entailed another decrease in the compartmentation.