Characterization of protein phosphorylation of the cytosol of AH-66 hepatoma ascites cells.
Endogenous phosphorylation reaction of the cytosol fraction of AH-66 hepatoma ascites cells in vitro was compared with that of normal rat liver. Cytosolic proteins with molecular weights of 125,000, 98,000, and 40,000 of AH-66 cells were heavily phosphorylated in a cyclic adenosine 3':5'-monophosphate-independent manner, but no counterpart was detected in normal liver cytosol. In order to examine whether these phosphoproteins were specifically present in AH-66 cytosol, cyclic adenosine 3':5'-monophosphate-independent protein kinases which phosphorylate these phosphoproteins were partially purified from AH-66 and liver cytosol by successive chromatography. Both kinase preparations were essentially free of endogenous protein substrates and catalyzed the phosphorylation of exogenous substrates, such as casein and phosvitin, but not histone and protamine. Both kinases markedly catalyzed the phosphorylation of the Mr 125,000, 98,000, and 40,000 proteins in AH-66 cytosol. The Mr 125,000, 98,000 and 40,000 proteins in liver cytosol were less intensely phosphorylated by the addition of the kinase from AH-66 cytosol. From these results, we conclude that these phosphoproteins are present in both AH-66 cytosol and liver cytosol but are highly concentrated in AH-66 cytosol.
D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008114:Liver Neoplasms, Experimental / Q000378:metabolism*; D008970:Molecular Weight; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification; D051381:Rats
D010750:Phosphoproteins; D000242:Cyclic AMP; D011494:Protein Kinases