Rat-liver cell sap protein was separated into four fractions by pH 5.5 precipitation and ammonium sulfate fractionation. On incubation with (32P)ATP, protein of two of the fractions was phosphorylated, showing the presence of protein kinase activity and endogenous protein substrates. The phosphorylation of one of the fractions was markedly stimulated by cyclic 3'5'AMP. Strong evidence was obtained that this phosphate-incorporating material was neither active phosphorylase, phosphorylase kinase nor glycogen synthetase. The two phosphate-incorporating fractions were chromatographed on DEAE-cellulose with stepwise elution. Several subtractions contained material which was phosphorylated on incubation with (32P)ATP. The phosphorylation of two subfractions was greatly stimulated by cyclic 3'5'AMP. By polyacrylamide gel electrophoresis in detergent of (32P)ATP-incubated material from these fractions, three major 32P-labelled components with estimated molecular weights of 62000, 55000 and 47 000 were demonstrated. It is supposed that these components are derived from enzymes or other proteins with functions regulated by phosphorylation-dephosphorylation reactions.
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