Breakdown of Na⁺/K⁺‐exchanging ATPase phosphoenzymes formed from ATP and from inorganic phosphate during Na⁺‐ATPase activity.

Abstract

The reactivity towards Na+ and K+ of Na+/K+-ATPase phosphoenzymes formed from ATP and Pi during Na+-ATPase turnover and that obtained from Pi in the absence of ATP, Na+ and K+ was studied. The phosphoenzyme formed from Pi in the absence of cycling and with no Na+ or K+ in the medium showed a biphasic time-dependent breakdown. The fast component, 96% of the total EP, had a decay rate of about 4 s(-1) in K+-free 130 mm Na+, and was 40% inhibited by 20 mm K+. The slow component, about 0.14 s(-1), was K+ insensitive. Values for the time-dependent breakdown of the phosphoenzymes obtained from ATP and from Pi during Na+-ATPase activity were indistinguishable from each other. In K+-free medium containing 130 mm Na+, the decays followed a single exponential with a rate constant of 0.45 s(-1). The addition of 20 mm K+ markedly increased the decays and made them biphasic. The fast components had a rate of approximately 220 s-1 and accounted for 92-93% of the total phosphoenzyme. The slow components decayed at a rate of about 47-53 s(-1). A second group of experiments examined the reactivity towards Na+ of the E2P forms obtained with ATP and Pi when the enzyme was cycling. In both cases, the rate of dephosphorylation was a biphasic function of [Na+]: inhibition at low [Na+], with a minimum at about 5 mm Na+, followed by recovery at higher [Na+]. Although qualitatively similar, the phosphoenzyme formed from Pi showed slightly less inhibition and more pronounced recovery. These results indicate that forward and backward phosphorylation during Na+-ATPase turnover share the same intermediates.

Frontal lobe bioenergetic metabolism in depressed adolescents with bipolar disorder: a phosphorus‐31 magnetic resonance spectroscopy study

“…Additional support for the relevance of Pi to the pathophysiology of BD is provided by the enduring observation that the activity of sodium–potassium adenosine triphosphatase (Na+/K+‐ATPase), an enzyme partially regulated by Pi (99–103), is altered in patients with BD (104, 105). In addition, there is a genetic association between BD and mutations in Na+/K+‐ATPase (106, 107).…”

Evidence for Tryptophan Residues in the Cation Transport Path of the Na+,K+-ATPase

“…Enzyme Phosphorylation by [γ-32 P]ATP. Phosphoenzyme formation was done manually for about 30 s at 0 °C (25). Aliquots of 50 µg of enzyme were incubated in a medium containing variable Na + concentrations (osmolarity was kept with choline), 30 mM imidazole (pH 7.5 at 0 °C), and 1 mM MgCl 2 with 10 µM [γ-32 P]ATP in a final volume of 100 µL.…”

Palytoxin and the sodium/potassium pump—phosphorylation and potassium interaction

“…Spontaneous transitions between both states, E1 and E2, may occur [27]; however, ionic pumping takes place due to transitions between substates of E1 and E2, according to chemical reactions of Na/K-ATPase with Na + , K + and ATP (table 1). The cycle, established by reactions 2-9, causes accumulation of extracellular Na + and intracellular K + , using the energy released by the ATP hydrolyses [28].…”

“…For intracellular ATP concentration in the order of 1 mM, the K + deocclusion in the intracellular space can be improved by the ATP binding [23,28]. In this case, first ATP binds to its low affinity site of the enzyme in the substate (K + ) 2 E2, forming the substate ATP low (K + ) 2 E2 (reaction 10).…”