Purification and characterization of protein kinase C from the nematode Caenorhabditis elegans.
Protein kinase C (PKC) of Caenorhabditis elegans was identified by enzymatic activity and [3H]phorbol 12,13-dibutyrate binding after DEAE-Sephacel column chromatography of a crude cytosolic extract. Ca(2+)-dependent activation of nematode PKC was observed in the presence of phosphatidylserine. The enzyme was maximally activated by 1,2-dioleoylglycerol or phorbol 12-myristate 13-acetate in the presence of phosphatidylserine and Ca2+. Hydroxyapatite column chromatography showed only one peak of PKC activity with histone H1 and myelin basic protein as substrates. The enzyme was purified to near homogeneity by sequential chromatography on polylysine-agarose and phosphatidylserine affinity columns. The purified protein showed a molecular mass of 79 kDa on SDS/PAGE. The substrate specificity of the C. elegans enzyme was shown to be different from that of mammalian PKCs. Here we describe some of the properties of the nematode enzyme.
Sassa
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NEC Fundamental Research Laboratories, Ibaraki, Japan.
Miwa
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J
D000818:Animals; D002107:Caenorhabditis / Q000201:enzymology*; D002122:Calcium Chloride / Q000494:pharmacology; D002845:Chromatography; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D004075:Diglycerides / Q000494:pharmacology; D017886:Durapatite; D006882:Hydroxyapatites; D007700:Kinetics; D008970:Molecular Weight; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D011493:Protein Kinase C / Q000302:isolation & purification* / Q000378:metabolism*; D013379:Substrate Specificity; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004075:Diglycerides; D006882:Hydroxyapatites; D015240:Phorbol 12,13-Dibutyrate; D017886:Durapatite; D011493:Protein Kinase C; D002122:Calcium Chloride; D013755:Tetradecanoylphorbol Acetate; C013965:diolein