title: Critical role of γ-phosphate in structural transition of Na,K-ATPase upon ATP binding.
abstract: Active transport of sodium and potassium ions by Na,K-ATPase is accompanied by the enzyme conformational transition between E1 and E2 states. ATP and ADP bind to Na,K-ATPase in the E1 conformation with similar affinity but the properties of enzyme in complexes with these nucleotides are different. We have studied thermodynamics of Na,K-ATPase binding with adenine nucleotides at different temperatures using isothermal titration calorimetry. Our data indicate that β-phosphate is involved in complex formation by increasing the affinity of adenine nucleotides to Na,K-ATPase by an order of magnitude, while γ-phosphate does not affect it. ATP binding to Na,K-ATPase in contrast to ADP binding generates a structural transition in the enzyme, which is consistent with the movement of a significant portion of the surface area to a solvent-protected state. We propose that ATP binding leads to convergence of the nucleotide-binding and phosphorylation domains transferring the enzyme from the "E1-open" to "E1-closed" conformation ready for phosphorylation.
authors.affiliation: 1] Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov St. 32, 119991 Moscow, Russia [2].
authors.forename: Irina Yu
authors.initials: IY
authors.lastname: Petrushanko
authors.affiliation: 1] Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov St. 32, 119991 Moscow, Russia [2].
authors.forename: Vladimir A
authors.initials: VA
authors.lastname: Mitkevich
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov St. 32, 119991 Moscow, Russia.
authors.forename: Anastasia A
authors.initials: AA
authors.lastname: Anashkina
authors.affiliation: Faculty of Biology, M.V. Lomonosov Moscow State University, 119234 Moscow, Russia.
authors.forename: Elizaveta A
authors.initials: EA
authors.lastname: Klimanova
authors.affiliation: Faculty of Biology, M.V. Lomonosov Moscow State University, 119234 Moscow, Russia.
authors.forename: Elena A
authors.initials: EA
authors.lastname: Dergousova
authors.affiliation: Faculty of Biology, M.V. Lomonosov Moscow State University, 119234 Moscow, Russia.
authors.forename: Olga D
authors.initials: OD
authors.lastname: Lopina
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov St. 32, 119991 Moscow, Russia.
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Makarov
mesh_terms: D000225:Adenine / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry*; D001665:Binding Sites; D003198:Computer Simulation; D004789:Enzyme Activation; D008956:Models, Chemical; D008958:Models, Molecular; D010710:Phosphates / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000648:ultrastructure*; D013696:Temperature
chemical_list: D010710:Phosphates; D000255:Adenosine Triphosphate; D000254:Sodium-Potassium-Exchanging ATPase; D000225:Adenine