Intact mitochondria prepared from pig heart muscle showed a considerable activity of creatine kinase(EC 2.7, 3.2), but this mitochindria bound creatine kinase was released in soluble form by incubation in isotonic sodium phosphate solution. This result suggests that the mitochondrial creatine kinase is rather loosely bound to the mitochondrial membrane outside.
The molecular nature and some of physico-chemical properties of mitochondrial creatine kinase were found to be different from that of cytoplasmic isoenzyme. The optimum pH in the direction of phosphocreatine formation is 8.2 for mitochondria bound creatine kinase ; 7.8, released and purified mitochondrial creatine kinase; 8.5, for cytoplasmic creatine kinase. The nature of substrate binding properties of the mitochondria bound creatine kinase(α=1) were found to be different from those of released mitochondrial creatine kinase(α=1/26) and cytoplasmic creatine kinase(α=1/3). The apparent Km values of the releasee mitochondrial creatine are 0.083 and 5.8 mM, repectively, and those of cytoplasmic creatine kinase, 0.34 and 12.8 mM. These kinetic properties indicate that mitochondrial creatine kinase is especially adapted to the formation of phosphocreatine.
The mitochondria bound creatine kinase was also found to be linked directly to the mitochondria respiratory system. The rate of respiration was controled by the amount of creatine and phosphocreatine added. The mitochondria respiration was also affected by the presence of cytoplasmic creatine kinase.
Based on the different nature of two forms of creatine kinase, a specific role of the mitochondrial creatine kinase in the heart cell energy metabolism was suggested.