Purification and characterization of two cyclic AMP-independent casein/glycogen synthase kinases from rat liver cytosol

Emilio Itarte, M. Angels Mor, Agustí Salavert, J. Manuel Pena, JoséFernando Bertomeu, Joan J. Guinovart

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Abstract

Two cyclic AMP-independent protein kinases (ATP: protein phosphotransferase, EC 2.7.1.37) (casein kinase 1 and 2) have been purified from rat liver cytosol by a method involving chromatography on phosphacellulose and casein-Sepharose 4B. Both kinases were essentially free of endogeneous protein substrates and capable of phosphorylating casein, phosvitin and I-form glycogen synthase, but were inactive on histone IIA, protamine and phosphorylase b. They were neither stimulated by cyclic AMP, Ca2+ and calmodulin, nor inhibited by the cyclic AMP-dependent protein kinase inhibitor protein. The casein and glycogen synthase kinase activities of each enzyme decreased at the same rate when incubated at 50°C. Casein kinase 1 and casein kinase 2 showed differences in molecular weight, sensitivity to KCl, Km for casein and phosvitin and Ka for Mg2+, whereas their Km values for ATP and I-form glycogen synthase were similar. The phosphorylation of glycogen synthase by these kinases correlated with a decrease in the ± glucose 6-phosphate activity ratio (independence ratio). However, casein kinase 1 catalyzed the incorporation of about 3.6 mol of 3-2-P 85 000 dalton subunit, decreasing the independence ratio from 83 to about 15, whereas the phosphorylation achieved by casein kinase 2 was only about 1.9 mol of 32P 85 000 dalton subunit, decreasing the independence ratio to about 23. The independence ratio decrease was prevented by the presence of casein but was unaffected by phosphorylase b. © 1981.
Original languageEnglish
Pages (from-to)334-347
JournalBBA - Enzymology
Volume658
DOIs
Publication statusPublished - 14 Apr 1981

Keywords

  • (Rat liver cytosol)
  • Casein kinase
  • Cyclic AMP
  • Glycogen synthase kinase
  • Protein kinase

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