Phosphorylation of pig epidermal soluble protein by endogenous cAMP-dependent protein kinase.
The distribution of adenosine 3',5'-monophosphate (cAMP)-dependent protein kinase and its substrate proteins was analyzed using soluble and particulate fractions of pig epidermal homogenates. When histone was used as a substrate for this enzyme reaction, protein kinase activity was distributed almost equally between the soluble and particulate fractions. However, the effect of exogenously added cAMP was confined almost exclusively to the soluble enzyme. Endogenous protein phosphorylation in the absence of exogenous histone was higher in the particulate fraction than in the soluble fraction, but the stimulating effect of cAMP was observed only in the soluble fraction. These results indicate that cAMP-dependent protein kinase is predominantly localized in the soluble fraction and phosphorylates soluble epidermal proteins. The particulate fraction contains protein kinase which is cAMP-independent and phosphorylates particulate-bound proteins as well as histone. Based on these observations, the soluble fraction was incubated with [gamma-32P]-ATP in the presence or absence of cAMP, and phosphorylated protein was analyzed by SDS disc- or slab-gel electrophoresis followed by autoradiography. Among many proteins whose phosphorylation was slightly increased by cAMP, a protein with Mr approximately 45,000 was found which was markedly phosphorylated in the presence of cAMP. Although this protein corresponds to one of the richest proteins in the epidermal soluble fraction, an important physiologic role for this phosphorylation has not been clarified.
D000818:Animals; D000242:Cyclic AMP / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D010750:Phosphoproteins / Q000302:isolation & purification; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D012867:Skin / Q000378:metabolism*; D012995:Solubility; D013552:Swine
D010750:Phosphoproteins; D011506:Proteins; D000242:Cyclic AMP; D011494:Protein Kinases