FLUXES THROUGH CYTOSOLIC AND MITOCHONDRIAL CREATINE-KINASE, MEASURED BY P-31 NMR

The kinetic properties of the cytoplasmic and the mitochondrial iso-en zymes of creatine kinase from striated muscle were studied in vitro an d in vivo. The creatine kinase (CK) iso-enzyme family has a multi-face ted role in cellular energy metabolism and is characterized by a compl ex pattern of tissue-specific expression and subcellular distribution. In mammalian tissues, there is al ways co-expression of at least two different CK isoforms. As a result, previous studies into the role of CK in energy metabolism have not been able to directly differentiate b etween the individual CK species. Here, we describe experiments which were directed at achieving this goal. First, we studied the kinetic pr operties of the muscle-specific cytoplasmic and mitochondrial CK isofo rms in purified form under in vitro, conditions, using a combination o f P-31 NMR and spectrophotometry. Secondly, P-31 NMR measurements of t he flux through the CK reaction were carried out on intact skeletal an d heart muscle from wild-type mice and from transgenic mice, homozygou s for a complete deficiency of the muscle-type cytoplasmic CK isoform. Skeletal muscle and heart were compared because they differ strongly in the relative abundance of the CK isoforms. The present data indicat e that the kinetic properties of cytoplasmic and mitochondrial CK are substantially different, both in vitro and in vivo. This finding parti cularly has implications for the interpretation of in vivo studies wit h P-31 NMR.