The Na-K-ATPase is characterized by a complex molecular heterogeneity that results from the expression and differential association of multiple isoforms of both its α- and β-subunits. At present, a...

Isozymes of the Na-K-ATPase: heterogeneity in structure, diversity in function

▪ Abstract The Na,K-ATPase or sodium pump carries out the coupled extrusion and uptake of Na and K ions across the plasma membranes of cells of most higher eukaryotes. It is a member of the P-type ...

Biochemistry of Na,K-ATPase

Structural organization, ion transport, and energy transduction of p-type atpases

Na+,K(+)-ATPase.

The sodium-potassium-activated adenosinetriphosphatase (Na(+)-K(+)-ATPase; Na(+)-K+ pump) is a ubiquitous plasma membrane enzyme that catalyzes the movement of K+ into cells in exchange for Na+. In addition, it provides the driving force for the transport of other solutes, notably amino acids, sugar, and phosphate. The regulation of Na(+)-K(+)-ATPase in various tissues is under the control of a number of circulating hormones that impart both short- and long-term control over its activity. The molecular mechanisms by which hormones alter Na(+)-K(+)-ATPase activity have only begun to be studied. In this review, we assess the acute and long-term actions of a number of hormones (aldosterone, thyroid hormone, catecholamines, insulin, carbachol) on the Na(+)-K+ pump. The long-term regulation exerted by thyroid hormone and aldosterone is mediated by changes in gene expression. The short-term regulation exerted by catecholamines is mediated by reversible phosphorylation of the pump catalytic subunit. Recent evidence supports regulation of the pump by phosphorylation in vitro and in intact cells. Finally, in some tissues the rapid action of insulin, aldosterone, and carbachol involves changes in the subcellular distribution of pump units.

Hormonal regulation of the Na(+)-K(+)-ATPase: mechanisms underlying rapid and sustained changes in pump activity

Synthesis and assembly of functional mammalian Na,K-ATPase in yeast.

The influence of beta subunit structure on the stability of Na+/K(+)-ATPase complexes and interaction with K+.

Recently, a beta subunit for the rat gastric H+,K(+)-ATPase (HK beta), which is structurally similar to the beta subunit of Na+, K(+)-ATPase, has been cloned and characterized. Using heterologous expression in yeast, we have tested the specificity of beta subunit assembly with different isoforms of the alpha subunit of Na+, K(+)-ATPase. Coexpression in yeast cells of the HK beta with both the sheep alpha 1 subunit and the rat alpha 3 subunit isoforms of Na+, K(+)-ATPase (alpha 1 and alpha 3, respectively) leads to the appearance of high-affinity ouabain-binding sites in yeast membranes. These ouabain-binding sites (alpha 1 plus HK beta, alpha 3 plus HK beta) have a high affinity for ouabain (Kd, 5-10 nM) and are expressed at levels similar to those formed with the rat beta 1 subunit of Na+, K(+)-ATPase (beta 1) (alpha 1 plus beta 1 or alpha 3 plus beta 1). Potassium acts as a specific antagonist of ouabain binding by alpha 1 plus HK beta and alpha 3 plus HK beta just like sodium pumps formed with beta 1. Sodium pumps formed with the HK beta, however, show quantitative differences in their affinity for ouabain and in the antagonism of K+ for ouabain binding. These data suggest that the structure of the beta subunit may play a role in sodium pump function.

http://www.pnas.org/content/89/7/2834.full.pdf

High-affinity ouabain binding by yeast cells expressing Na+, K(+)-ATPase alpha subunits and the gastric H+, K(+)-ATPase beta subunit.

The Influence of Subunit Structure on the Interaction of Na/K-ATPase Complexes with Na.: A CHIMERIC β SUBUNIT REDUCES THE Na DEPENDENCE OF PHOSPHOENZYME FORMATION FROM ATP

Amino acids N886-A911 of the rat Na+-K+-ATPase alpha3-subunit were replaced by the corresponding region (Q905-V930) of the rat gastric H+-K+-ATPase alpha-subunit. The chimera (NGH26) was expressed in yeast with the rat Na+-K+ -ATPase beta1-subunit (rbeta1), the rat H+-K+-ATPase beta-subunit (HKbeta), the chimeric beta-subunit NHbeta1 (containing the carboxy-terminal ectodomain of HKbeta), or the chimeric beta-subunit HNbeta1 (containing the carboxy-terminal ectodomain of rbeta1). Increased resistance to trypsin digestion indicated that NGH26 preferentially assembled with HKbeta and NHbeta1 rather than with rbeta1 or HNbeta1. Ouabain binding also indicated that more functional complexes were assembled when NGH26 was expressed with HKbeta or NHbeta1. These results suggest that the sequence Q905-V930 interacts with the HKbeta-subunit on the extracellular side of the cell membrane. The NGH26 + HKbeta complex is less stable than alpha3 + HKbeta when heated and also has a lower binding affinity for ouabain [dissociation constant (Kd) = 63 nM] compared with alpha3 + rbeta1 or alpha3 + HKbeta (K(d) = 5-10 nM). In contrast, the NGH26+NHbeta1 complex is thermally as stable as alpha3 + rbeta1 complexes, and its ouabain binding affinity (K(d) = 10 nM) is the same as the wild type. These results indicate that the amino acids Q905-V930 of the rat gastric H+-K+-ATPase alpha-subunit preferentially associate with the extracellular domain of H+-K+-ATPase beta-subunit to form functional pump complexes and that the cytoplasmic and/or transmembrane region of the beta-subunit influences the stability of the alpha beta complexes.

Kurt A. Eakle

Papers

Synthesis and assembly of functional mammalian Na,K-ATPase in yeast.

The influence of beta subunit structure on the stability of Na+/K(+)-ATPase complexes and interaction with K+.

High-affinity ouabain binding by yeast cells expressing Na+, K(+)-ATPase alpha subunits and the gastric H+, K(+)-ATPase beta subunit.

The Influence of Subunit Structure on the Interaction of Na/K-ATPase Complexes with Na.: A CHIMERIC β SUBUNIT REDUCES THE Na DEPENDENCE OF PHOSPHOENZYME FORMATION FROM ATP

Na+-K+-ATPase alpha-subunit containing Q905-V930 of gastric H+-K+-ATPase alpha preferentially assembles with H+-K+-ATPase beta.