Cytochrome c and Ouabain Binding Site of Na,K-ATPase

Na,K-ATPase is an electrogenic pump found in cell plasma membranes that acts as the basic unit of animal life. This enzyme is highly susceptible to cardiotonic steroid (CTS) inhibition. The role of Na,K-ATPase in signaling has introduced a novel viewpoint regarding the enzyme's function, as the ouabain-binding site is involved in several physiological processes. At high concentrations, ouabain blocks Na⁺ and K⁺ ion transport by Na,K-ATPase, whereas at low concentrations, it activates the signaling function of the enzyme. Notably, Na,K-ATPase does not fit into the categories of G protein-coupled receptors or ligand-gated ion channels. This indicates that it may be a distinct cell surface receptor that interacts with signaling molecules through allosteric regulation. In the present study, we have identified new modulators of Na,K-ATPase sensitivity to ouabain, and studied the kinetic effects of physiological concentrations of ouabain on Na,K-ATPase in the hippocampus. Specifically, Cytochrome c (Cytc) increases an affinity for ouabain and the maximal velocity (Vmax) of the enzyme. After binding to Na,K-ATPase, ouabain induces conformational changes that drive shifts between enzymatic cycles.