Structure of Supramolecular Clusters of Creatine Kinases in the Intermembrane Space of Mitochondria

The structure of rat-heart mitochondria and mitochondrial membrane preparations is studied using cryoelectron tomography. It is found that pseudocrystalline clusters formed by densely packed proteins exist between the inner and outer membranes. Using subtomogram averaging, the structure of these clusters is obtained and it is shown that they are formed by tens or even hundreds of densely packed octamers of mitochondrial creatine kinases (mCK). In intact mitochondria, mCK octamers form a single membrane-parallel layer in which they are tightly coupled to the outer and inner membranes, as well as to each other. On membrane fragments in which there is no compression of the intermembrane space, the supramolecular mCK clusters are organized differently: they form two layers in which there are fewer contacts with the membrane, but more protein–protein contacts. In addition, bilayer clusters generally lack curvature and are larger in size, which is indicative of their increased rigidity and possible role in stabilizing the membrane when it is damaged. The obtained results are discussed in the context of previously known data on the functional role, structure, and location of creatine kinases in mitochondria.