Hilbers F et al. (2016), Tuning of the Na,K-ATPase by the beta subunit.

XB-ART-52945

Sci Rep 2016 Feb 05;6:20442. doi: 10.1038/srep20442.

Show Gene links Show Anatomy links

Tuning of the Na,K-ATPase by the beta subunit.

Hilbers F , Kopec W , Isaksen TJ , Holm TH , Lykke-Hartmann K , Nissen P , Khandelia H , Poulsen H .

???displayArticle.abstract???
The vital gradients of Na(+) and K(+) across the plasma membrane of animal cells are maintained by the Na,K-ATPase, an αβ enzyme complex, whose α subunit carries out the ion transport and ATP hydrolysis. The specific roles of the β subunit isoforms are less clear, though β2 is essential for motor physiology in mammals. Here, we show that compared to β1 and β3, β2 stabilizes the Na(+)-occluded E1P state relative to the outward-open E2P state, and that the effect is mediated by its transmembrane domain. Molecular dynamics simulations further demonstrate that the tilt angle of the β transmembrane helix correlates with its functional effect, suggesting that the relative orientation of β modulates ion binding at the α subunit. β2 is primarily expressed in granule neurons and glomeruli in the cerebellum, and we propose that its unique functional characteristics are important to respond appropriately to the cerebellar Na(+) and K(+) gradients.

???displayArticle.pubmedLink??? 26847162
???displayArticle.pmcLink??? PMC4742777
???displayArticle.link??? Sci Rep

Species referenced: Xenopus laevis
Genes referenced: atp1a1 rbfox3

???attribute.lit??? ???displayArticles.show???

	Figure 1. Electrophysiological properties of Î±1Î²1 and Î±1Î²2.A simplified Post-Albers scheme with Na+ in purple and K+ in green is shown in (a). Difference curves in K+-free buffer with and without 10âmM ouabain for (b) Î±1Î²1,(c) Î±1Î²2 are shown. The curves were fitted with single exponentials, giving the voltage dependent (d) charge translocation from the off currents and (e) rate constants from the on currents. Nâ=â3â10 with oocytes from at least two Xenopus laevis females. Data are represented as meanâÂ±âSD.
	Figure 2. Expression and localization of the Î²1 and Î²2 isoforms in mouse brain.(a) Western blot analysis of the indicated isolated mouse brain regions using antibodies against Î²2 and GAPDH. (b) Fluorescence immunohistochemistry of Î²1 and Î²2 in cerebellum co-stained with the neuronal marker NeuN. GCL: granule cell layer. P: Purkinje cell layer. ML: molecular layer. Scale bars represent 20âÎ¼m.
	Figure 3. Charge translocation curves of chimeras and pocket mutants.Charge translocation was determined for Î±1 coexpressed with (a) Î² chimeras, where the N-terminal, the C-terminal or the transmembrane region of Î²1 was replaced with that of Î²2, with (b) Î² mutants where smaller stretches in the transmembrane region of Î²1 were replaced with the corresponding Î²2 sequences, FK with AF N-terminally, AGI with TAM in the middle or the C-terminal 16 residues, or (c) a combination of the FK toAF , AGI to TAM and VSD to ISE (at the C-terminus, cf. Fig. S6) in Î²1 giving Î²1/3mut. Nââ¥â5 with oocytes from at least two Xenopus laevis females. Data are represented as meanâÂ±âSD.
	Figure 4. Molecular dynamics simulations of Î±1Î²1 and Î±1Î²2.(a) Atomistic models of the Î± (white-grey) and Î²1 (blue) subunits, embedded in the POPC lipid membrane (contour shown in magenta). Several important residues of the Î² subunit are shown in spacefill. Note that 33â34 FK and 61â63 ISE residues are located at or near the membrane interface. Important residues forming ion binding sites I, II and III are shown as sticks, and bound sodium ions are shown as pink spheres. Water and the Î³ subunit are omitted for clarity, but included in the model. (b) Comparison of the helix tilt between the transmembrane helix of Î²1 (blue) and Î²2 (light green). The tilt is defined as the angle between the helix axis and the z-axis, which is perpendicular to the membrane surface. Presented values are the averages from the last 40âns, with error estimations obtained with block averaging. (c,d) Interaction patterns between the Î² helix (blue Î²1 C) and light green Î²2 D)) and the M7 helix (yellow) and the C-terminus (orange) of the Î± subunit in Î±1Î²1 C) and Î±1Î²2 D). Interactions between selected residues (shown in licorice) are shown as purple springs, with minimum distances recorded in simulations (last 40âns) indicated in italics. Hydrophobic carbon atoms are shown in cyan, oxygen atoms bearing partial negative charge are shown in red and nitrogen atoms bearing partial positive charge are shown in deep blue.
	Figure 5. Flexibility of the Î± C-terminus with Î²1 or Î²2.(a) Root mean square deviation (RMSD) of the C-terminus of Î±1 with Î²1 or Î²2 showing displacement in Î±1Î²2. (b) Root mean square fluctuations (RMSF) of heavy atom positions of the protein residues that form the C-terminal tail of Î±1, average of the last 60âns of the MD trajectory.

References [+] :

Albers, Biochemical aspects of active transport. 1967, Pubmed