Ouabain Binding Kinetics of the Rat Alpha Two and Alpha Three Isoforms of the Sodium-Potassium Adenosine Triphosphate

O’Brien, William J.; Lingrel, Jerry B.; Wallick, Earl T.

doi:10.1006/abbi.1994.1136

Cited by 154 publications

(111 citation statements)

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“…6A, Fig. S4A, and Movie S2), which selectively blocks rodent NAKα3 activity (26). Unlike ASPD and ouabain, staurosporine, an apoptosis inducer that inhibits multiple kinases, induced shrinkage but not swelling (Fig.…”

Section: Aspd Bind To Nakα3mentioning

confidence: 97%

Na, K-ATPase α3 is a death target of Alzheimer patient amyloid-β assembly

Ohnishi

Yanazawa

Sasahara

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

116

155

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Neurodegeneration correlates with Alzheimer's disease (AD) symptoms, but the molecular identities of pathogenic amyloid β-protein (Aβ) oligomers and their targets, leading to neurodegeneration, remain unclear. Amylospheroids (ASPD) are AD patient-derived 10-to 15-nm spherical Aβ oligomers that cause selective degeneration of mature neurons. Here, we show that the ASPD target is neuronspecific Na + /K + -ATPase α3 subunit (NAKα3). ASPD-binding to NAKα3 impaired NAKα3-specific activity, activated N-type voltage-gated calcium channels, and caused mitochondrial calcium dyshomeostasis, tau abnormalities, and neurodegeneration. NMR and molecular modeling studies suggested that spherical ASPD contain N-terminal-Aβ-derived "thorns" responsible for target binding, which are distinct from low molecular-weight oligomers and dodecamers. The fourth extracellular loop (Ex4) region of NAKα3 encompassing Asn 879 and Trp 880 is essential for ASPD-NAKα3 interaction, because tetrapeptides mimicking this Ex4 region bound to the ASPD surface and blocked ASPD neurotoxicity. Our findings open up new possibilities for knowledge-based design of peptidomimetics that inhibit neurodegeneration in AD by blocking aberrant ASPD-NAKα3 interaction.NMR | computational modeling | abnormal protein-protein interaction in synapse | hyperexcitotoxicity | protein-protein interaction inhibitors

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Section: Aspd Bind To Nakα3mentioning

confidence: 97%

Na, K-ATPase α3 is a death target of Alzheimer patient amyloid-β assembly

Ohnishi

Yanazawa

Sasahara

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

116

155

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“…In humans and most other mammals, ␣ 1 has high affinity for ouabain (low nanomolar K d ), but in rodents ␣ 1 has a 100-to 1,000-fold lower ouabain affinity (84). Most cells also express either ␣ 2 or ␣ 3 Na ϩ pumps (sperm express ␣ 4 ) and, in contrast to ␣ 1 , the high ouabain affinity of ␣ 2 and ␣ 3 (K d s in the low nanomolar range) (84) has been conserved through mammalian evolution (90).…”

Section: How Does Low-dose Ouabain Augment Contractility?mentioning

confidence: 99%

How does salt retention raise blood pressure?

Blaustein¹,

Zhang²,

Ling³

et al. 2006

American Journal of Physiology-Regulatory, Integrative and Comp

129

121

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A critical question in hypertension research is: How is long-term blood pressure controlled? Excessive NaCl ingestion or NaCl retention by the kidneys and the consequent tendency toward plasma volume expansion lead to hypertension. Nevertheless, the precise mechanisms linking salt to high blood pressure are unresolved. The discovery of endogenous ouabain, an adrenocortical hormone, provided an important clue. Ouabain, a selective Na ϩ pump inhibitor, has cardiotonic and vasotonic effects. Plasma endogenous ouabain levels are significantly elevated in Ϸ40% of patients with essential hypertension and in animals with several forms of salt-dependent hypertension. Also, prolonged ouabain administration induces hypertension in rodents. Mice with mutant Na ϩ pumps or Na/Ca exchangers (NCX) and studies with a ouabain antagonist and an NCX blocker are revealing the missing molecular mechanisms. These data demonstrate that ␣ 2 Na ϩ pumps and NCX1 participate in long-term regulation of vascular tone and blood pressure. Pharmacological agents or mutations in the ␣ 2 Na ϩ pump that interfere with the action of ouabain on the pump, and reduced NCX1 expression or agents that block NCX all impede the development of salt-dependent or ouabain-induced hypertension. Conversely, nanomolar ouabain, reduced ␣ 2 Na ϩ pump expression, and smooth muscle-specific overexpression of NCX1 all induce hypertension. Furthermore, ouabain and reduced ␣ 2 Na ϩ pump expression increase myogenic tone in isolated mesenteric small arteries in vitro, thereby tying these effects directly to the elevation of blood pressure. Thus, endogenous ouabain, and vascular ␣ 2 Na ϩ pumps and NCX1, are critical links between salt and hypertension. New pharmacological agents that act on these molecular links have potential in the clinical management of hypertension.ouabain; Na ϩ pump; Na/Ca exchanger; Ca 2ϩ ; myogenic tone HYPERTENSION, DEFINED AS A diastolic blood pressure (BP) Ն 90 mmHg and/or systolic BP Ն 140 mmHg, is endemic in Westernized societies. This is a very important public health issue because hypertension is a major risk factor for premature death and disability from heart attack, heart failure, stroke, and many other afflictions (16,59). In the United States, alone, Ϸ20% of the population (i.e., Ϸ50 million individuals) are hypertensive; moreover, more than half of all individuals over the age of 60 years have hypertension. In a small fraction of cases, the hypertension is due to specific causes, such as renal vascular disease or excessive secretion of aldosterone (primary aldosteronism) or catecholamines (pheochromocytoma). The vast majority (Ϸ90%) of patients, however, have elevated BP of unknown cause; hence the terms, primary or essential hypertension. The immediate cause for the elevated BP in nearly all chronic hypertensive persons is excessive narrowing of the small (resistance) arteries. Nevertheless, a key question in any discussion of hypertension is: What specific mechanisms actually lead to the abnormal arterial constriction and elevation ...

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“…It contains the binding and phosphorylation ATP domain, as well as essential amino acids for the binding of sodium, potassium and some inhibitors (8,16,17). The ß subunit has a structural function and is important for membrane enzyme insertion (8,14,18,19). The g peptide has hydrophobic characteristics associated with the Na,K-ATPase molecule; however, it is not required for the enzyme catalytic activity or for ion transport.…”

Section: Introductionmentioning

confidence: 99%

Solubilization of Na,K-ATPase from rabbit kidney outer medulla using only C12E8

Santos

Lamas

Ciancaglini

2002

Braz J Med Biol Res

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SDS, C 12 E 8 , CHAPS or CHAPSO or a combination of two of these detergents is generally used for the solubilization of Na,K-ATPase and other ATPases. Our method using only C 12 E 8 has the advantage of considerable reduction of the time for enzyme purification, with rapid solubilization and purification in a single chromatographic step. Na,KATPase-rich membrane fragments of rabbit kidney outer medulla were obtained without adding SDS. Optimum conditions for solubilization were obtained at 4ºC after rapid mixing of 1 mg of membrane Na,K-ATPase with 1 mg of C 12 E 8 /ml, yielding 98% recovery of the activity. The solubilized enzyme was purified by gel filtration on a Sepharose 6B column at 4ºC. Non-denaturing PAGE revealed a single protein band with phosphomonohydrolase activity. The molecular mass of the purified enzyme estimated by gel filtration chromatography was 320 kDa. The optimum apparent pH obtained for the purified enzyme was 7.5 for both PNPP and ATP. The dependence of ATPase activity on ATP concentration showed high (K 0.5 = 4.0 µM) and low (K 0.5 = 1.4 mM) affinity sites for ATP, with negative cooperativity. Ouabain (5 mM), oligomycin (1 µg/ml) and sodium vanadate (3 µM) inhibited the ATPase activity of C 12 E 8 -solubilized and purified Na,KATPase by 99, 81 and 98.5%, respectively. We have shown that Na,KATPase solubilized only with C 12 E 8 can be purified and retains its activity. The activity is consistent with the form of (aß) 2 association.

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Ouabain Binding Kinetics of the Rat Alpha Two and Alpha Three Isoforms of the Sodium-Potassium Adenosine Triphosphate

Cited by 154 publications

References 0 publications

Na, K-ATPase α3 is a death target of Alzheimer patient amyloid-β assembly

Na, K-ATPase α3 is a death target of Alzheimer patient amyloid-β assembly

How does salt retention raise blood pressure?

Solubilization of Na,K-ATPase from rabbit kidney outer medulla using only C12E8

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