Download MendeleyStructures and characterization of digoxin- and bufalin-bound Na+,K+-ATPase compared with the ouabain-bound complex.
Laursen, M., Gregersen, J.L., Yatime, L., Nissen, P., Fedosova, N.U.(2015) Proc Natl Acad Sci U S A 112: 1755-1760
- PubMed: 25624492
- DOI: https://doi.org/10.1073/pnas.1422997112
- Primary Citation of Related Structures:
4RES, 4RET - PubMed Abstract:
Cardiotonic steroids (CTSs) are specific and potent inhibitors of the Na(+),K(+)-ATPase, with highest affinity to the phosphoenzyme (E2P) forms. CTSs are comprised of a steroid core, which can be glycosylated, and a varying number of substituents, including a five- or six-membered lactone. These functionalities have specific influence on the binding properties. We report crystal structures of the Na(+),K(+)-ATPase in the E2P form in complex with bufalin (a nonglycosylated CTS with a six-membered lactone) and digoxin (a trisaccharide-conjugated CTS with a five-membered lactone) and compare their characteristics and binding kinetics with the previously described E2P-ouabain complex to derive specific details and the general mechanism of CTS binding and inhibition. CTSs block the extracellular cation exchange pathway, and cation-binding sites I and II are differently occupied: A single Mg(2+) is bound in site II of the digoxin and ouabain complexes, whereas both sites are occupied by K(+) in the E2P-bufalin complex. In all complexes, ¦ÁM4 adopts a wound form, characteristic for the E2P state and favorable for high-affinity CTS binding. We conclude that the occupants of the cation-binding site and the type of the lactone substituent determine the arrangement of ¦ÁM4 and hypothesize that winding/unwinding of ¦ÁM4 represents a trigger for high-affinity CTS binding. We find that the level of glycosylation affects the depth of CTS binding and that the steroid core substituents fine tune the configuration of transmembrane helices ¦ÁM1-2.
Organizational Affiliation:
Centre for Membrane Pumps in Cells and Disease, Danish National Research Foundation, DK-8000 Aarhus C, Denmark; Departments of Biomedicine and.
