Adenosine 3',5'-monophosphate-dependent protein kinase in supernatants from dispersed bovine parathyroid cells.
The presence and characteristics of protein kinase(s) were studied in supernatants of sonicates of dispersed bovine parathyroid cells. cAMP caused a 3- to 5-fold stimulation of protein kinase activity in such extracts, with half of the maximal activation at 4--5 x 10(-8) M cAMP. Protein kinase inhibitor nearly totally abolished both basal and cAMP-stimulated activity, suggesting that most of the activity was cAMP dependent. About 90% of the cAMP-stimulated protein kinase activity eluted from a DEAE-cellulose column at 0.15 m NaCl, consistent with a type II enzyme. The presence of a type II enzyme was also supported by the effects of histone and salt concentrations on enzyme activity; both the basal and cAMP-stimulated activity ratios (activity minus cAMP divided by activity plus 10(-6) M cAMP) were stable in 0.4 M NaCl. The basal activity ratio was not increased by concentrations of histone as high as 10 mg/ml in the protein kinase assay. The predominance of the type II enzyme in dispersed bovine parathyroid cells made it possible to develop conditions for extracting the enzyme from intact intact cells (0.4 m NaCl and 5 mg/ml charcoal), whereby the state of activation of the enzyme remained relatively constant. These studies demonstrate the presence of cAMP-dependent protein kinase activity in dispersed bovine parathyroid cells and define conditions which make it possible to assess the effects of various secretagogues on protein kinase activation in intact parathyroid cells.
D000818:Animals; D002417:Cattle; D002606:Charcoal; D002852:Chromatography, Ion Exchange; D000242:Cyclic AMP / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D010280:Parathyroid Glands / Q000201:enzymology*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
D047428:Protein Kinase Inhibitors; D002606:Charcoal; D012965:Sodium Chloride; D000242:Cyclic AMP; D011494:Protein Kinases