%0 Journal Article
%A Fruergaard, Marlene U.
%A Dach, Ingrid
%A Andersen, Jacob L.
%A Ozol, Mette
%A Shahsavar, Azadeh
%A Quistgaard, Esben M.
%A Poulsen, Hanne
%A Fedosova, Natalya U.
%A Nissen, Poul
%T The Na<sup>+</sup>,K<sup>+</sup>-ATPase in complex with beryllium fluoride mimics an ATPase phosphorylated state 
%J The journal of biological chemistry
%V 298
%N 9
%@ 0021-9258
%C Bethesda, MD.
%I American Soc. for Biochemistry and Molecular Biology
%M PUBDB-2022-07058
%P 102317
%D 2022
%Z ISSN 0021-9258 not unique: **2 hits**.
%X The Na<sup>+</sup>,K<sup>+</sup>-ATPase generates electrochemical gradients of Na<sup>+</sup> and K<sup>+</sup> across the plasma membrane via a functional cycle that includes various phosphoenzyme intermediates. However, the structure and function of these intermediates and how metal fluorides mimick them require further investigation. Here, we describe a 4.0 Å resolution crystal structure and functional properties of the pig kidney Na<sup>+</sup>,K<sup>+</sup>-ATPase stabilized by the inhibitor beryllium fluoride (denoted E2–BeF<sub>x</sub>). E2–BeF<sub>x</sub> is expected to mimic properties of the E2P phosphoenzyme, yet with unknown characteristics of ion and ligand binding. The structure resembles the E2P form obtained by phosphorylation from inorganic phosphate (P<sub>i</sub>) and stabilized by cardiotonic steroids, including a low-affinity Mg<sup>2+</sup> site near ion binding site II. Our anomalous Fourier analysis of the crystals soaked in Rb<sup>+</sup> (a K<sup>+</sup> congener) followed by a low-resolution rigid-body refinement (6.9–7.5 Å) revealed preocclusion transitions leading to activation of the dephosphorylation reaction. We show that the Mg<sup>2+</sup> location indicates a site of initial K<sup>+</sup> recognition and acceptance upon binding to the outward-open E2P state after Na<sup>+</sup> release. Furthermore, using binding and activity studies, we find that the BeF<sub>x</sub>-inhibited enzyme is also able to bind ADP/ATP and Na<sup>+</sup>. These results relate the E2–BeF<sub>x</sub> complex to a transient K<sup>+</sup>- and ADP-sensitive E∗P intermediate of the functional cycle of the Na<sup>+</sup>,K<sup>+</sup>-ATPase, prior to E2P.
%F PUB:(DE-HGF)16
%9 Journal Article
%$ pmid:35926706
%U <Go to ISI:>//WOS:000863323200005
%R 10.1016/j.jbc.2022.102317
%U https://bib-pubdb1.desy.de/record/486058