3.A.3.1.1
Na⁺-, K⁺-ATPase (Na⁺ efflux; K⁺ uptake).  Kinetic alterations due to a missense mutation in the alpha2 subunit cause familial hemiplegic migraine type 2 (Segall et al. 2004). Mutation in the γ-subunit causes renal hypomagnesemia, associated with hypocalciurea (Cairo et al., 2008). The Na/K-ATPase is an important signal transducer that not only interacts and regulates protein kinases, but also functions as a scaffold (Li and Xie, 2009). Capsazepine, a synthetic vanilloid, converts the Na, K-ATPase to a Na-ATPase (Mahmmoud, 2008a). There are alternative α- and β-subunits, α₁, α₂,... β₁, β_2,... in muscle which form α₁β₁, α₁β₂, α₂β₁ and α₂β₂, heterodimers, each with differing Na⁺ affinities (4-13mM) (Kristensen and Juel, 2010). α₃ and β₃ isoforms have also been identified. The γ-subunit is the same as TC# 1.A.27.2.1. Poulsen et al. (2010) have proposed a second ion conduction pathway in the C-terminal part of the ATPase. The two C-terminal tyrosines stabilize the occluded Na/K pump conformations containing Na or K ions (Vedovato and Gadsby, 2010). Na⁺, K⁺-ATPase mutations causing familial hemiplegic migraines type 2 (FHM2) inhibit phosphorylation (Schack et al., 2012). Salt, the vascular Na⁺/K⁺ ATPase and the endogenous glycosides, ouabain and marinobufagenin, play roles in systemic hypertension (Hauck and Frishman, 2012). Protein kinase A (PKA) phosphorylation of Ser936 (in the intracellular loop between transmembrane segments M8 and M9) opens an intracellular C-terminal water pathway leading to the third Na⁺-binding site (Poulsen et al., 2012). PKA-mediated phosphorylation regulates activity in vivo. Ser-938 is located (Einholm et al. 2016). E960 on the Na⁺-K⁺-ATPase and F28 on phospholemman (PLM) are critical for phospholemman (PLM) inhibition, but there is at least one additional site that is important for tethering PLM to the ATPase. Mutations in the Na⁺/K⁺-ATPase α3 subunit gene (ATP1A3) cause rapid-onset dystonia-parkinsonism, a rare movement disorder characterized by sudden onset of dystonic spasms and slow movements (Doğanli et al. 2013).  The 3-d strcuture of the Na⁺-bound Na⁺,K⁺-ATPase at 4.3 Å resolution reveals the positions of the three Na⁺ ions (Nyblom et al. 2013).  Mutations cause adrenal hypertension (Kopec et al. 2014) as well as alternating hemiplegia of childhood (AHC) and rapid-onset dystonia- parkinsonism (RDP) (Rosewich et al. 2014).  Differences in the structures of the ouabain-, digonxin- and bufalin-bound enzyme have been reported (Laursen et al. 2015).  ATPase inhibitors have been shown to be effective anti-cancer agents (Alevizopoulos et al. 2014). Cys45 in the β-subunit can be glutathionylated, regulating the activity of the enzyme (Garcia et al. 2015). ATP1A2 mutations play a role in migraine headaches (Friedrich et al. 2016). The beta2 subunit is essential for motor physiology in mammals, and in contrast to beta1 and beta3, beta2 stabilizes the Na⁺-occluded E1P state relative to the outward-open E2P state (Hilbers et al. 2016). Numerous transcription factors, hormones, growth factors, lipids, and extracellular stimuli as well as epigenetic signals modulate the transcription of Na,K-ATPase subunits (Li and Langhans 2015). Čechová et al. 2016 have identified two cytoplasmic pathways along the pairs of TMSs, TMS3/TMS7 or TM6S/TMS9 that allow hydration of the cation binding sites or transport of cations from/to the bulk medium. Dissipation of the transmembrane gradient of K⁺ and Na⁺ due to ouabain inhibition increases Ptgs2 and Nr4a1 transcription by increasing Ca²⁺ influx through L-type Ca²⁺ channels that, in turn, leads to CaMKII-mediated phosphorylation of CREB and calcineurin-mediated dephosphorylation of NFAT, respectively (Smolyaninova et al. 2017). ZMay play a role in the development of gastric adenocarcinomas (Wang et al. 2017). Mutations F785L and T618M give rise to familial rapid onset dystonia parkonsonism by distinct mechanisms (Rodacker et al. 2006). Reacts with methylglyoxal to inhibit its activity (Svrckova et al. 2017).  Accumulation of beta-amyloid (Abeta) at the early stages of Alzheimer's disease is accompanied by reduction of Na,K-ATPase functional activity. Petrushanko et al. 2016 showed that monomeric Abeta(1-42) forms a tight (Kd of 3 mμM), enthalpy-driven equimolar complex with alpha1beta1 Na,K-ATPase. Complex formation results in dose-dependent inhibition of the enzyme hydrolytic activity. The binding site of Abeta(1-42) is localized in the "gap" between the α- and β-subunits of Na,K-ATPase, disrupting the enzyme functionality by preventing the subunits from shifting towards each other. Interaction of Na,K-ATPase with exogenous Abeta(1-42) leads to a pronounced decrease of the enzyme transport and hydrolytic activities and Src-kinase activation in neuroblastoma cells SH-SY5Y. This interaction allows regulation of Na,K-ATPase activity by short-term increases in the Abeta(1-42) level (Petrushanko et al. 2016). Two distinct phospholipids bind to two distinct sites on the ATPase, affecting activity and stability (Habeck et al. 2017). Five cysteinyl residues (C452, C456, C457, C577, and C656) serve as the cisplatin binding sites on the cytoplasmic loop connecting transmembrane helices 4 and 5 (Šeflová et al. 2018). Mutations can cause F/SHM with moderate penitrance (Prontera et al. 2018).  Arginine substitution of a cysteine in transmembrane helix M8 converts the Na⁺,K⁺-ATPase to an electroneutral pump similar to the gastric H⁺,K⁺-ATPase (Holm et al. 2017). Early onset life-threatening epilepsy can be associated with ATP1A3 gene variants (Ishihara et al. 2019), and loss of Na/K pump function is the common feature of mutants that induce hyperaldosteronism (Meyer et al. 2019). Three Na⁺ sites are defined in the Na⁺-bound crystal structure of the Na⁺, K⁺-ATPase. Sites I and II overlap with two K⁺ sites in the K⁺-bound structure, whereas site III is unique and Na⁺ specific. A glutamine in transmembrane helix M8 (Q925) appears from the crystal structures to coordinate Na⁺ at site III, but does not contribute to K⁺ coordination at sites I and II. Nielsen et al. 2019 addressed the functional role of Q925 in the various conformational states of this-ATPase by examining the mutants Q925A/G/E/N/L/I/Y both enzymatically and electrophysiologically, thereby revealing their Na⁺ and K⁺ binding properties. Q925 substitutions had minor effects on Na⁺ binding from the intracellular side of the membrane, but mutations Q925A and Q925G increased the apparent Na⁺ affinity, but caused dramatic reductions of the binding of K⁺ as well as Na⁺ from the extracellular side of the membrane. Thus, an interaction between sites III and I and a possible gating function of Q925 in the release of Na⁺ at the extracellular side are supported (Nielsen et al. 2019). The alpha2 Na⁺/K⁺-ATPase isoform mediates LPS-induced neuroinflammation (Leite et al. 2020). Apical periodontitis induces changes on oxidative stress parameters and increases Na⁺/K⁺-ATPase activity in adult rats (Barcelos et al. 2020). Familial hemiplegic migraine type 2 can be due to a missense mutation (L425H) in ATP1A2 (Antonaci et al. 2021). Mutations in ATP1A3 and FXYD genes (TC# 1.A.27) can cause childhood-onset schizophrenia (Chaumette et al. 2020). The ATPase plays a role in cell adhesion, motility, and migration of cancer cells (Silva et al. 2021). TMS2 moves outward as Na⁺ is deoccluded from the E1 conformation (Young and Artigas 2021). Kinetic properties and crystal structures of the Na⁺,K⁺-ATPase in complex with cardiotonic steroids (CTS) has revealed differences between CTS subfamilies. Ladefoged et al. 2021 found beneficial effects of K⁺ on bufadienolide binding in contrast to the well-known antagonism between K⁺ and cardenolides. Bufadienolide binding is affected by (i) electrostatic attraction of the lactone ring by a cation and (ii) the ability of a cation to stabilize and "shape" the site constituted by transmembrane helices of the alpha-subunit (αM1-6). The latter effect was due to varying coordination patterns involving amino acid residues from helix bundles αM1-4 and αM5-10. Substituents on the steroid core of a bufadienolide add to and modify the cation effects (Ladefoged et al. 2021). The Na⁺-K⁺-ATPase functions in the developing hippocampus (Shao et al. 2021). CryoEM analyses of the Na⁺,K⁺-ATPase in the two E2P states with and without cardiotonic steroids has revealed mechanistic details (Kanai et al. 2022). A conserved ion-binding site tyrosine plays a role in ion selectivity of the Na⁺/K⁺ pump (Spontarelli et al. 2022). Essential roles of the Na+K+-ATPase in ischemic pathology provide a platform for the improvement in clinical research on ischemic stroke (Zhu et al. 2022). In intact live cells, the regulatory complex is composed of two alpha subunits associated with two beta subunits, decorated with two PLM regulatory subunits. Docking and molecular dynamics (MD) simulations generated a structural model of the complex. alpha-alpha subunit interactions support conformational coupling of the catalytic subunits, which may enhance the NKA turnover rate (Seflova et al. 2022). There is a phenotypic continuum of ATP1A3-related disorders (Vezyroglou et al. 2022). The Na⁺/K⁺-ATPase is central to the pathogenesis of neurological diseases such as alternating hemiplegia of childhood. This ATPase has 3 distinct ion binding sites I-III. Binding of Na⁺ at each site in the human alpha3 Na⁺/K⁺-ATPase can be resolved using extracellular Na⁺-mediated transient currents. When the ATPase is constrained to bind and release only Na⁺, three kinetic components: fast, medium, and slow, can be isolated, depending on the voltage step direction and the occlusion (or deocclusion) of each of the 3 Na⁺s. Patient-derived mutations of residues which coordinate Na⁺ at site III exclusively impact the slow component, demonstrating that site III is crucial for deocclusion and release of the first Na⁺ into the extracellular milieu (Moreno et al. 2022). The opening of ion channels in cardiomyocytes is regulated by the surface electric double layer of the cell membrane as revealed by studies with digoxin (Zhou et al. 2022). The effects of H₂O₂ on cys residues and other targets in the Na,K-ATPase have been discussed (Chkadua et al. 2022). The influence of the Na⁺/K⁺-ATPase on neurodegenerative diseases has been reviewed (Zhang et al. 2022). The presence of two oppositely directed transmembrane ion gradients (for Na⁺ and K⁺) is important for robust stabilization of cellular volume in human erythrocytes (Ataullakhanov et al. 2022).  This ATPase plays important roles in neurodegenerative diseases (Zhang et al. 2022). Na/K-ATPase signaling tonically inhibits sodium reabsorption in the renal proximal tubule (Mukherji et al. 2023). Hypoxic stress-dependent regulation of Na,K-ATPase in ischemic heart disease has been reviewed (Baloglu 2023).  Chronic testosterone deficiency increases late inward sodium current and promotes triggered activity in ventricular myocytes from aging male mice (Banga et al. 2023). Three dynamic lipid interaction sites in the plasma membrane Na⁺,K⁺-ATPase influence activity of the transporter (Mahato and Andersson 2023). Interaction of pumps and transporters positioned at distant biological membranes with various forms of energy transfer between them may result in hypoxic/reperfusion injury, different kinds of muscle fatigue, and nerve-glia interactions (Dimitrov 2023). Marinobufagenin (MBG) is a member of the bufadienolide family of compounds, which are natural cardiac glycosides found in a variety of animal species, including man, which have different physiological and biochemical functions but have a common action on the inhibition of the Na⁺/K⁺-ATPase. MBG acts as an endogenous cardiotonic steroid, and in the last decade, its role as a pathogenic factor in various human diseases has emerged (Carullo et al. 2023).  A malfunctioning gene product is required for disease induction by ATP1A1 variants and that if a pathology is associated with protein-null variants, they may display low penetrance or high age of onset (Spontarelli et al. 2023). The Na,K-ATPase is a murzyme, facilitating thermodynamic equilibriums at the membrane-interface (Manoj et al. 2023).  The Na⁺K⁺-ATPase is present in boar sperm HPM, and it changes during capacitation (Awda et al. 2023). Clinical features of CAPOS syndrome are caused by maternal ATP1A3 gene variation (Gao et al. 2024).Inositol hexakisphosphate kinase 1 (IP6K1), governs the degradation of Na⁺/K⁺-ATPase via an autoinhibitory domain of PI3K p85α (Jin et al. 2024).  Two lysines in the transmembrane segments contribute to generate a pump with reduced stoichiometry (2 K⁺/1 Na⁺), allowing Artemia to maintain steeper Na⁺ gradients in hypersaline environments (Artigas et al. 2023).  Na⁺/K⁺-ATPase participates in Ca²⁺-signaling transduction and neurotransmitter release by coordinating the ion gradient across the cell membrane. This ATPase works synergistically with ion channels to form a dynamic network of ion homeostatic regulation and affects cellular communication (Huang et al. 2024).  Bufalin is a direct anticancer drug and a Na⁺/K⁺-ATPase inhibitor by forcing the Na⁺/Ca²⁺ exchanger to reverse its function, which transfers Ca²⁺ into the cytoplasm and ultimately causes Ca²⁺ overload-triggered pyroptosis (Li et al. 2024). Proximal tubule angiotensin II signaling regulates Na⁺ transporters in the mouse nephron (Prontera et al. 2018).  Arginine substitution of a cysteine in transmembrane helix M8 converts the Na⁺,K⁺-ATPase to an electroneutral pump similar to the gastric H⁺,K⁺-ATPase (Holm et al. 2017). Early onset life-threatening epilepsy can be associated with ATP1A3 gene variants (Ishihara et al. 2019), and loss of Na/K pump function is the common feature of mutants that induce hyperaldosteronism (Meyer et al. 2019). Three Na⁺ sites are defined in the Na⁺-bound crystal structure of the Na⁺, K⁺-ATPase. Sites I and II overlap with two K⁺ sites in the K⁺-bound structure, whereas site III is unique and Na⁺ specific. A glutamine in transmembrane helix M8 (Q925) appears from the crystal structures to coordinate Na⁺ at site III, but does not contribute to K⁺ coordination at sites I and II. Nielsen et al. 2019 addressed the functional role of Q925 in the various conformational states of this-ATPase by examining the mutants Q925A/G/E/N/L/I/Y both enzymatically and electrophysiologically, thereby revealing their Na⁺ and K⁺ binding properties. Q925 substitutions had minor effects on Na⁺ binding from the intracellular side of the membrane, but mutations Q925A and Q925G increased the apparent Na⁺ affinity, but caused dramatic reductions of the binding of K⁺ as well as Na⁺ from the extracellular side of the membrane. Thus, an interaction between sites III and I and a possible gating function of Q925 in the release of Na⁺ at the extracellular side are supported (Nielsen et al. 2019). The alpha2 Na⁺/K⁺-ATPase isoform mediates LPS-induced neuroinflammation (Leite et al. 2020). Apical periodontitis induces changes on oxidative stress parameters and increases Na⁺/K⁺-ATPase activity in adult rats (Barcelos et al. 2020). Familial hemiplegic migraine type 2 can be due to a missense mutation (L425H) in ATP1A2 (Antonaci et al. 2021). Mutations in ATP1A3 and FXYD genes (TC# 1.A.27) can cause childhood-onset schizophrenia (Chaumette et al. 2020). The ATPase plays a role in cell adhesion, motility, and migration of cancer cells (Silva et al. 2021). TMS2 moves outward as Na⁺ is deoccluded from the E1 conformation (Young and Artigas 2021). Kinetic properties and crystal structures of the Na⁺,K⁺-ATPase in complex with cardiotonic steroids (CTS) has revealed differences between CTS subfamilies. Ladefoged et al. 2021 found beneficial effects of K⁺ on bufadienolide binding in contrast to the well-known antagonism between K⁺ and cardenolides. Bufadienolide binding is affected by (i) electrostatic attraction of the lactone ring by a cation and (ii) the ability of a cation to stabilize and "shape" the site constituted by transmembrane helices of the alpha-subunit (αM1-6). The latter effect was due to varying coordination patterns involving amino acid residues from helix bundles αM1-4 and αM5-10. Substituents on the steroid core of a bufadienolide add to and modify the cation effects (Ladefoged et al. 2021). The Na⁺-K⁺-ATPase functions in the developing hippocampus (Shao et al. 2021). CryoEM analyses of the Na⁺,K⁺-ATPase in the two E2P states with and without cardiotonic steroids has revealed mechanistic details (Kanai et al. 2022). A conserved ion-binding site tyrosine plays a role in ion selectivity of the Na⁺/K⁺ pump (Spontarelli et al. 2022). Essential roles of the Na+K+-ATPase in ischemic pathology provide a platform for the improvement in clinical research on ischemic stroke (Zhu et al. 2022). In intact live cells, the regulatory complex is composed of two alpha subunits associated with two beta subunits, decorated with two PLM regulatory subunits. Docking and molecular dynamics (MD) simulations generated a structural model of the complex. alpha-alpha subunit interactions support conformational coupling of the catalytic subunits, which may enhance the NKA turnover rate (Seflova et al. 2022). There is a phenotypic continuum of ATP1A3-related disorders (Vezyroglou et al. 2022). The Na⁺/K⁺-ATPase is central to the pathogenesis of neurological diseases such as alternating hemiplegia of childhood. This ATPase has 3 distinct ion binding sites I-III. Binding of Na⁺ at each site in the human alpha3 Na⁺/K⁺-ATPase can be resolved using extracellular Na⁺-mediated transient currents. When the ATPase is constrained to bind and release only Na⁺, three kinetic components: fast, medium, and slow, can be isolated, depending on the voltage step direction and the occlusion (or deocclusion) of each of the 3 Na⁺s. Patient-derived mutations of residues which coordinate Na⁺ at site III exclusively impact the slow component, demonstrating that site III is crucial for deocclusion and release of the first Na⁺ into the extracellular milieu (Moreno et al. 2022). The opening of ion channels in cardiomyocytes is regulated by the surface electric double layer of the cell membrane as revealed by studies with digoxin (Zhou et al. 2022). The effects of H₂O₂ on cys residues and other targets in the Na,K-ATPase have been discussed (Chkadua et al. 2022). The influence of the Na⁺/K⁺-ATPase on neurodegenerative diseases has been reviewed (Zhang et al. 2022). The presence of two oppositely directed transmembrane ion gradients (for Na⁺ and K⁺) is important for robust stabilization of cellular volume in human erythrocytes (Ataullakhanov et al. 2022).  This ATPase plays important roles in neurodegenerative diseases (Zhang et al. 2022). Na/K-ATPase signaling tonically inhibits sodium reabsorption in the renal proximal tubule (Mukherji et al. 2023). Hypoxic stress-dependent regulation of Na,K-ATPase in ischemic heart disease has been reviewed (Baloglu 2023).  Chronic testosterone deficiency increases late inward sodium current and promotes triggered activity in ventricular myocytes from aging male mice (Banga et al. 2023). Three dynamic lipid interaction sites in the plasma membrane Na⁺,K⁺-ATPase influence activity of the transporter (Mahato and Andersson 2023). Interaction of pumps and transporters positioned at distant biological membranes with various forms of energy transfer between them may result in hypoxic/reperfusion injury, different kinds of muscle fatigue, and nerve-glia interactions (Dimitrov 2023). Marinobufagenin (MBG) is a member of the bufadienolide family of compounds, which are natural cardiac glycosides found in a variety of animal species, including man, which have different physiological and biochemical functions but have a common action on the inhibition of the Na⁺/K⁺-ATPase. MBG acts as an endogenous cardiotonic steroid, and in the last decade, its role as a pathogenic factor in various human diseases has emerged (Carullo et al. 2023).  A malfunctioning gene product is required for disease induction by ATP1A1 variants and that if a pathology is associated with protein-null variants, they may display low penetrance or high age of onset (Spontarelli et al. 2023). The Na,K-ATPase is a murzyme, facilitating thermodynamic equilibriums at the membrane-interface (Manoj et al. 2023).  The Na⁺K⁺-ATPase is present in boar sperm HPM, and it changes during capacitation (Awda et al. 2023). Clinical features of CAPOS syndrome are caused by maternal ATP1A3 gene variation (Gao et al. 2024).Inositol hexakisphosphate kinase 1 (IP6K1), governs the degradation of Na⁺/K⁺-ATPase via an autoinhibitory domain of PI3K p85α (Jin et al. 2024).  Two lysines in the transmembrane segments contribute to generate a pump with reduced stoichiometry (2 K⁺/1 Na⁺), allowing Artemia to maintain steeper Na⁺ gradients in hypersaline environments (Artigas et al. 2023).  Na⁺/K⁺-ATPase participates in Ca²⁺-signaling transduction and neurotransmitter release by coordinating the ion gradient across the cell membrane. This ATPase works synergistically with ion channels to form a dynamic network of ion homeostatic regulation and affects cellular communication (Huang et al. 2024).  Bufalin is a direct anticancer drug and a Na⁺/K⁺-ATPase inhibitor by forcing the Na⁺/Ca²⁺ exchanger to reverse its function, which transfers Ca²⁺ into the cytoplasm and ultimately causes Ca²⁺ overload-triggered pyroptosis (Li et al. 2024). Proximal tubule angiotensin II signaling regulates Na⁺ transporters in the mouse nephron (Nelson et al. 2021).