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Phosphorylation of the α-Subunit of Na,K-ATPase from Duck Salt Glands by cAMP-Dependent Protein Kinase Inhibits the Enzyme Activity

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Abstract

Although it was shown earlier that phosphorylation of Na,K-ATPase by cAMP-dependent protein kinase (PKA) occurs in intact cells, the purified enzyme in vitro is phosphorylated by PKA only after treatment by detergent. This is accompanied by an unfortunate side effect of the detergent that results in complete loss of Na,K-ATPase activity. To reveal the effect of Na,K-ATPase phosphorylation by PKA on the enzyme activity in vitro, the effects of different detergents and ligands on the stoichiometry of the phosphorylation and activity of Na,K-ATPase from duck salt glands (α1β1-isoenzyme) were comparatively studied. Chaps was shown to cause the least inhibition of the enzyme. In the presence of 0.4% Chaps at 1 : 10 protein/detergent ratio in medium containing 100 mM KCl and 0.3 mM ATP, PKA phosphorylates serine residue(s) of the Na,K-ATPase with stoichiometry 0.6 mol Pi/mol of α-subunit. Phosphorylation of Na,K-ATPase by PKA in the presence of the detergent inhibits the Na,K-ATPase. A correlation was found between the inclusion of Pi into the α-subunit and the loss of activity of the Na,K-ATPase.

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Authors and Affiliations

  1. Department of Biochemistry and 2 Department of Bioorganic Chemistry, School of Biology, Lomonosov Moscow State University, Moscow, 119899, Russia

    D. A. Murtazina, A. M. Rubtsov & O. D. Lopina

  2. Department of Biochemistry and Department of Chemistry, Carleton University, Ottawa KIS 5B6, Canada

    K. B. Storey

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  1. D. A. Murtazina
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  2. S. P. Petukhov
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  3. A. M. Rubtsov
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  4. K. B. Storey
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  5. O. D. Lopina
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Murtazina, D.A., Petukhov, S.P., Rubtsov, A.M. et al. Phosphorylation of the α-Subunit of Na,K-ATPase from Duck Salt Glands by cAMP-Dependent Protein Kinase Inhibits the Enzyme Activity. Biochemistry (Moscow) 66, 865–874 (2001). https://doi.org/10.1023/A:1011900718655

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