The mitochondria prepared from swine heart muscle tissues showed a considerable degree of the activity of membrane bound creatine kinase. The enzyme was found to be bound loosely to the mitochondrial membrane, and thus the most of the bound enzyme could be released from the mitochondria by treating with isotonic sodium phosphate buffer solution at neutral pH. The mitochondrial enzyme exhibited its characteristic properties different from those of the cytoplasmic enzyme : their substrate affinities for creatine and phosphoryl creatine were found to be different each others. In addition, it was demonstrated that the bound enzyme was engaged in the mitochondrial respiratory system. In the heart muscle tissue system, phosphoryl creating generated by mitochondrial creatine kinase is the end product of oxidative phosphorylation in the presence of physiological concentrations of creatine and ADP. Thus phosphoryl creatine may take an important role of the control of mitochondrial respiration by the ratio of phosphoryl creatine. Based on the results obtained, it was suggested that membrane bound creatine kinase may have a specific function for the transfer of high energy phosphate bond generated by the mitochondrial respiratory system to the energy utilizing sites.