Purification and Characterization of Human Sarcomeric Mitochondrial Creatine Kinase

In order to set the basis for detailed clinical investigations, mitochondrial creatine kinase (Mi-CK) was purified to homogeneity from human cardiac muscle. Biophysical characterization by SDS-PAGE, gel permeation chromatography and by electron microscopy of negatively stained single molecules demonstrated that, similar to other vertebrate Mi-CKs, human sarcomeric Mi-CK occurs in two different oligomeric forms, dimers and octamers, that are readily interconvertible. The apparentMrs of Mi-CK protomers, dimers and octamers were 43 600±800, 79 700±800 and 371 000±3 000, respectively. In addition, isoelectric focussing proved to be a suitable technique for routinely distinguishing Mi-CK from cytosolic MM-CK and gave pIvalues of 8.30±0.04 and 7.44±0.04 for octameric and dimeric human sarcomeric Mi-CK. Circumstantial evidence suggests that both the highly symmetrical structure and the high pIvalue of Mi-CK octamers are crucial determinants for the physiological functions of this enzyme.