New protein kinase from plasma membrane of Ehrlich ascites tumor cells activated by natural polypeptides.
A polypeptide-dependent protein kinase was purified about 80-fold from an extract of plasma membranes of Ehrlich ascites tumor cells. The membranes were extracted with Nonidet P-40, and the extract was purified by ammonium sulfate fractionation and hydroxylapatite and affinity chromatography. The activity was stimulated 10-fold or more by polypeptide preparations from a variety of tissues, including placenta and hypothalamus. Polypeptide-dependent protein kinase had a pH optimum of about 7.5 and required Mg2+ for activity. Mn2+ at low concentrations (200 microM) stimulated enzyme activity somewhat but inhibited activity strongly at higher concentrations. The best available substrate for polypeptide-dependent protein kinase was beta-casein, and little or no phosphorylation was observed with alpha-casein, kappa-casein, phosvitin, alpha-lactalbumin, alpha-lactoglobulin, and histone. However, several endogenous substrates from plasma membranes of Ehrlich ascites tumor cells were phosphorylated. Polypeptide-dependent protein kinase activity was not inhibited by 10 mM N-ethylmaleimide, and this resistance was useful in differentiating this protein kinase from other protein kinases that were present in crude fractions and sensitive to the inhibitor.
D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000201:enzymology*; D002364:Caseins / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D007031:Hypothalamus / Q000032:analysis; D007768:Lactalbumin / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D008297:Male; D051379:Mice; D010455:Peptides / Q000494:pharmacology*; D010774:Phosvitin / Q000378:metabolism; D010920:Placenta / Q000032:analysis; D011247:Pregnancy; D011494:Protein Kinases / Q000378:metabolism*; D013379:Substrate Specificity
D002364:Caseins; D010455:Peptides; D010774:Phosvitin; D007768:Lactalbumin; D011494:Protein Kinases; D008274:Magnesium