Distribution and characterization of cAMP-dependent protein kinase isoenzymes in bovine adrenal cells.
Subcellular localization and characterization of cAMP-kinase isoenzymes in fasciculata reticularis bovine adrenal cells has been investigated. Different subcellular fractions were purified on a Percoll gradient and characterized by marker enzymes. cAMP-kinase was located principally in cytosol and microsomes. In the low-speed particulate fractions cAMP-kinase was found associated mainly with plasma membrane but not with mitochondria. Characterization of isoenzyme patterns in subcellular fractions by conventional DEAE-cellulose chromatography and by anion-exchange HPLC gives essentially the same results. Isoenzyme I appears to be the main enzyme in cytosol whereas isoenzyme II predominates in solubilized microsome and plasma membrane enriched fraction. Photoaffinity labelling of chromatographic fractions demonstrated that HPLC separates both cAMP binding subunits. Photoaffinity labelling of the different subcellular fraction by 8-azido-[32P]cAMP confirmed the data obtained by anion-exchange chromatography. However, in microsomes this method revealed the presence of both isoenzymes and the preferential solubilization of isoenzyme II by Triton X-100. In summary, our results indicate a subcellular compartmentalization of cAMP-kinase in bovine adrenal cells with a preferential localization of isoenzyme I in cytosol and of isoenzyme II in membrane. However, the relation between the distribution and the role of each isoenzyme has so far not been documented.
D000311:Adrenal Glands / Q000201:enzymology* / Q000378:metabolism; D000345:Affinity Labels; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D002499:Centrifugation, Density Gradient; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology; D008928:Mitochondria / Q000201:enzymology; D010777:Photochemistry; D011494:Protein Kinases / Q000378:metabolism*; D012995:Solubility
D000345:Affinity Labels; D007527:Isoenzymes; D000242:Cyclic AMP; D011494:Protein Kinases