Immunological quantitation of phospholipid/Ca2+-dependent protein kinase and its fragments. Tissue levels, subcellular distribution, and ontogenetic changes in brain and heart.
Levels of phospholipid/Ca2+-dependent protein kinase (protein kinase C, 80 kDa) and its presumed proteolytic fragments were quantified in a variety of animal tissues and cultured human leukemic cell lines (HL60 and K562) using an immunoblot analysis technique. Of many tissues examined, the rat brain and HL60 cells were by far the richest sources of the 80-kDa native enzyme, with its concentration estimated to be about 2-3 microM in both tissues. The major enzyme species detected in most tissues, however, was the 67-kDa fragment; the 80-kDa native enzyme was present in a smaller amount except in spleen which contained nearly equal amounts of both enzyme species. It was also found that HL60 and K562 leukemic cells contained the 50-kDa species instead of the 67-kDa species. A study of the subcellular distribution of the 80- and 67-kDa species showed the enzyme to be localized predominantly in the soluble fraction for some tissues (e.g. heart) and nearly equally distributed between soluble and particulate fractions in others (e.g. spleen). In the brain, however, the majority of the enzyme was present in the particulate fraction, in agreement with the findings made with immunocytochemical localization of the enzyme. The total enzyme content in developing rat brain and heart increased during the first 2 to 4 weeks following birth and decreased to 60% of peak levels in the adult. The present immunological method, showing for the first time that the tissue levels of phospholipid/Ca2+-dependent protein kinase and its fragments can be quantitated, would be useful for studies on the regulation of cellular events and pathophysiology of tissues thought to be involved in this multi-functional protein phosphorylation system.
D000818:Animals; D001345:Autoradiography; D001921:Brain / Q000201:enzymology* / Q000254:growth & development; D004591:Electrophoresis, Polyacrylamide Gel; D006321:Heart / Q000254:growth & development; D007158:Immunologic Techniques; D009206:Myocardium / Q000201:enzymology*; D010446:Peptide Fragments / Q000032:analysis*; D011493:Protein Kinase C / Q000032:analysis*; D051381:Rats; D013347:Subcellular Fractions / Q000201:enzymology; D014018:Tissue Distribution
D010446:Peptide Fragments; D011493:Protein Kinase C