Two methods to avoid the effect of endogenous protein inhibitors during the assay of protein kinase C activity in tissue extracts.
Using H1 as substrate the protein kinase C activity of rat liver cell sap was increased about fourfold by treatment with DEAE-cellulose at pH 7.5 at an intermediate ionic strength due to removal of protein inhibitors. The activity of cell sap from rat spleen, brain or muscle was about doubled by the same treatment. In contrast, when a specific synthetic peptide substrate was used the corresponding increase of enzyme activity was not obtained when the inhibitors were removed. This shows that this type of substrates should be preferred for reliable assays of protein kinase C in crude extracts. The possible role of the protein inhibitors for the substrate specificity of protein kinase C is briefly discussed.
Ekman
P
P
Department of Medical and Physiological Chemistry, Uppsala University, Sweden.
Eller
M
M
Ragnarsson
U
U
Engström
L
L
D000818:Animals; D008099:Liver / Q000737:chemistry; D008722:Methods; D010761:Phosphorus Radioisotopes; D011493:Protein Kinase C / Q000032:analysis*; D011506:Proteins / Q000037:antagonists & inhibitors*; D051381:Rats; D013154:Spleen / Q000737:chemistry; D013379:Substrate Specificity; D013552:Swine; D014020:Tissue Extracts / Q000737:chemistry*
D010761:Phosphorus Radioisotopes; D011506:Proteins; D014020:Tissue Extracts; D011493:Protein Kinase C