title: Structural insights into the high affinity binding of cardiotonic steroids to the Na+,K+-ATPase.
abstract: The Na+,K+-ATPase belongs to the P-ATPase family, whose characteristic property is the formation of a phosphorylated intermediate. The enzyme is also a defined target for cardiotonic steroids which inhibit its functional activity and initiate intracellular signaling. Here we describe the 4.6 Å resolution crystal structure of the pig kidney Na+,K+-ATPase in its phosphorylated form stabilized by high affinity binding of the cardiotonic steroid ouabain. The steroid binds to a site formed at transmembrane segments αM1-αM6, plugging the ion pathway from the extracellular side. This structure differs from the previously reported low affinity complex with potassium. Most importantly, the A domain has rotated in response to phosphorylation and αM1-2 move towards the ouabain molecule, providing for high affinity interactions and closing the ion pathway from the extracellular side. The observed re-arrangements of the Na+,K+-ATPase stabilized by cardiotonic steroids may affect protein-protein interactions within the intracellular signal transduction networks.
authors.affiliation: Centre for Membrane Pumps in Cells and Disease-PUMPKIN, Danish National Research Foundation, Denmark.
authors.forename: Laure
authors.initials: L
authors.lastname: Yatime
authors.forename: Mette
authors.initials: M
authors.lastname: Laursen
authors.forename: J Preben
authors.initials: JP
authors.lastname: Morth
authors.forename: Mikael
authors.initials: M
authors.lastname: Esmann
authors.forename: Poul
authors.initials: P
authors.lastname: Nissen
authors.forename: Natalya U
authors.initials: NU
authors.lastname: Fedosova
mesh_terms: D000818:Animals; D001665:Binding Sites; D002316:Cardiotonic Agents / Q000737:chemistry*; D018360:Crystallography, X-Ray; D008274:Magnesium / Q000737:chemistry; D008958:Models, Molecular; D010042:Ouabain / Q000737:chemistry*; D010766:Phosphorylation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors* / Q000737:chemistry; D013552:Swine
chemical_list: D002316:Cardiotonic Agents; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D008274:Magnesium