Purification and isotype analysis of protein kinase C from rat liver nuclei.
The properties and subtype composition of protein kinase C present in rat liver nuclei were studied in a Triton-X-100 extract of isolated purified nuclei. The enzyme activity was dependent on both Ca2+ and phosphatidylserine, but the phorbol ester 12-O-tetradecanoylphorbol 13-acetate gave only a partial stimulation. Both histone and myelin basic protein served as substrate. Purification of the Triton-X-100 extract followed by Q-Sepharose chromatography gave a preparation with a specific activity of 70 pmol/mg protein min. Western blotting of this preparation showed only the presence of the delta and zeta subtypes, but not the alpha-subtype, although the latter was present in rat liver homogenates. The beta, gamma and epsilon subtypes were not found in the homogenate nor in the nuclear extract. The specific activity of protein kinase C could be further increased up to 800 pmol/mg protein min after protamine agarose chromatography. Also in this preparation the presence of the delta and zeta subtypes could be established.
de Moel
M P
MP
Department of Biochemistry, University of Nijmegen, The Netherlands.
Van Emst-De Vries
S E
SE
Willems
P H
PH
De Pont
J J
JJ
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002467:Cell Nucleus / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification*; D008099:Liver / Q000201:enzymology*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D017830:Octoxynol; D010718:Phosphatidylserines / Q000494:pharmacology; D011493:Protein Kinase C / Q000737:chemistry / Q000302:isolation & purification*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007527:Isoenzymes; D010718:Phosphatidylserines; D017830:Octoxynol; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium