Direct interaction of beta-amyloid with Na,K-ATPase as a putative regulator of the enzyme function

Abstract

By maintaining the Na+ and K+ transmembrane gradient mammalian Na,K-ATPase acts as a key regulator of neuronal electrotonic properties. Na,K-ATPase has an important role in synaptic transmission and memory formation. Accumulation of beta-amyloid (Aβ) at the early stages of Alzheimer’s disease is accompanied by reduction of Na,K-ATPase functional activity. The molecular mechanism behind this phenomenon is not known. Here we show that the monomeric Aβ(1-42) forms a tight (Kd of 3 μM), enthalpy-driven equimolar complex with α1β1 Na,K-ATPase. The complex formation results in dose-dependent inhibition of the enzyme hydrolytic activity. The binding site of Aβ(1-42) is localized in the “gap” between the alpha- and beta-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 Aβ(1-42) leads to a pronounced decrease of the enzyme transport and hydrolytic activity and Src-kinase activation in neuroblastoma cells SH-SY5Y. This interaction allows regulation of Na,K-ATPase activity by short-term increase of the Aβ(1-42) level. However prolonged increase of Aβ(1-42) level under pathological conditions could lead to chronical inhibition of Na,K-ATPase and disruption of neuronal function. Taken together, our data suggest the role of beta-amyloid as a novel physiological regulator of Na,K-ATPase.

Phosphorylation of the Amyloid-Beta Peptide Inhibits Zinc-Dependent Aggregation, Prevents Na,K-ATPase Inhibition, and Reduces Cerebral Plaque Deposition

“…The purified preparation of Na + /K + -ATPase (α1β1 isozyme) was obtained from duck salt glands as described elsewhere ( Petrushanko et al, 2016 ). The purity grade of Na + /K + -ATPase was 99% and specific activity of the enzyme reached ∼2400 μmol of Pi (mg of protein × h) −1 at 37°C.…”

“…Hydrolytic activity of Na + /K + -ATPase in the purified preparation was measured as ouabain-sensitive (1 mM) ATP cleavage in the reaction medium containing 130 mM NaCl, 20 mM KCl, 3 mM MgCl 2 , 3 mM ATP, and 30 mM imidazole, pH 7.4, 37°C as elsewhere ( Petrushanko et al, 2016 ). Stock solutions containing Aβ 42 and pS8-Aβ 42 peptides, prepared as described above, were added to the reaction medium (without ATP) up to a concentration of 40 μM.…”

“…Previously, it was shown that Na + ,K + -ATPase activity was inhibited in post-mortem tissues of AD patients and in amyloid-containing hippocampi of transgenic mice (but not in the amyloid-free cerebellum) ( Dickey et al, 2005 ; Kreutz et al, 2013 ; Zhang et al, 2013 ). The latest studies demonstrated that Aβ 42 in form of monomers or oligomers directly binds to Na + ,K + -ATPase, which results in the inhibition of the enzyme as well as the triggering of intracellular signaling cascades ( Ohnishi et al, 2015 ; Petrushanko et al, 2016 ). Therefore, phosphorylation at Ser8 might neutralize some pathogenic properties of Aβ.…”

Regulation of Neuronal Na+/K+-ATPase by Specific Protein Kinases and Protein Phosphatases

“…Partial NKA inhibition or NKA deficiency in brain has also been implicated in spatial learning and memory deficits (Sato et al, 2004;Moseley et al, 2007), as well as in the pathophysiology of Alzheimer's disease (e.g., Ohnishi et al, 2015;Petrushanko et al, 2016). Thus, the ability of activated PKG and PKC to reversibly inhibit neuronal NKAs suggests that these kinases may be downstream effectors of signaling pathways that couple primary messengers to NKAs, through which multiple brain functions can be regulated or dysregulated.…”

Na+/K+-pump and neurotransmitter membrane receptors

“…In contrast, sub-µM angiotensin-(1-7) activates Na + /K + -ATPase from rat cerebral cortex synaptosomal membranes (Lopez Ordieres et al 1998 ). β -Amyloid has also been proposed as a regulator of Na + /K + -ATPase (Petrushanko et al 2016 ). These authors found that β -amyloid inhibited Na + /K + -ATPase activity of neuroblastoma cells by binding between the α and β subunits of the enzyme and preventing the subunits moving towards each other.…”