Beneficent and maleficent effects of cations on bufadienolide binding to Na+,K+-ATPase

Kinetic properties and crystal structures of the Na+,K+-ATPase in complex with cardiotonic steroids (CTS) revealed significant differences between CTS subfamilies (Laursen et al., 2015): beneficial effects of K+ on bufadienolide binding strongly contrasted with K+/cardenolide antagonism. To solve this riddle we applied docking and molecular dynamics simulations of the complexes involving Na+,K+-ATPase, bufadienolides (bufalin, cinobufagin), and ions (K+, Na+, Mg2+). The results revealed that bufadienolide binding is affected by i) electrostatic attraction of the lactone ring by a cation, and ii) the ability of a cation to stabilize and "shape" the site constituted by transmembrane helices of the -subunit (M1-6). The latter effect was due to varying coordination patterns involving amino acid residues from helix bundles M1-4 and M5-10. Substituents on the steroid core of a bufadienolide add to and modify the cation effects. The above rationale is fully consistent with the ion effects on the kinetics of Na+,K+-ATPase/bufadienolide interactions.