Properties of a protein kinase C activity in synaptic plasma membrane and postsynaptic density fractions isolated from canine cerebral cortex.
Protein kinase C (PKC) activity (phosphorylation increased by addition of Ca2+/phosphatidylserine or Ca2+/phosphatidylserine/phorbol ester) was found in both a synaptic plasma membrane (SPM) and a postsynaptic density (PSD) fraction. The SPM fraction had as endogenous substrates 87K-, 60K-, 50K-, and 20K-Mr proteins, whereas the PSD fraction had only the 20K-Mr protein. The PKC activity was also detected using histone III-S as a substrate, in SPM but much less in PSD. Phosphorylations of histone and the endogenous substrates of PKC, assayed in the absence of Ca2+, were enhanced in the SPM prepared after treatment of brain homogenate with phorbol 12-myristate 13-acetate (TPA), but very little enhancement was found in PSD after such treatment. The SPM PKC activity (both for endogenous substrate proteins and for histone), which was enhanced by TPA treatment of brain homogenate, was inhibited by calcium (IC50, 3 x 10(-7) M). The phosphorylations of the 20K-Mr protein in PSD, and in SPM prepared with and without TPA treatment, were all inhibited by H-7. The 20K-Mr protein in the PSD fraction is also phosphorylated by a PSD Ca2+/calmodulin-dependent protein kinase II. The evidence indicates that both SPM and PSD fractions contain a PKC activity. Detergent treatment of SPM, to produce a purified PSD fraction, results in a PSD fraction that has lost most of the endogenous substrates, lost the TPA-induced enhanced activity assayed in the absence of Ca2+, and lost the inhibitory effect of low Ca2+ concentration.
Suzuki
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Laboratory of Cell Biology, Rockefeller University, New York, New York 10021.
Siekevitz
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P
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000494:pharmacology; D002540:Cerebral Cortex / Q000201:enzymology*; D004285:Dogs; D004533:Egtazic Acid / Q000494:pharmacology; D007700:Kinetics; D010718:Phosphatidylserines / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D013569:Synapses / Q000201:enzymology*; D013570:Synaptic Membranes / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002147:Calmodulin; D010718:Phosphatidylserines; D004533:Egtazic Acid; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium