Rapid purification of protein kinase C by high performance liquid chromatography

Protein kinase C was purified from rat brain cytosol by using a high performance liquid chromatography (HPLC), Pharmacia FPLC system. This procedure employed a column chromatography on DE-52, followed by three steps of HPLC procedures with threonine-Sepharose (prepared as described in this report), TSK gel Phenyl-5PW (Toyo Soda), and TSK gel G3000SW (Toyo Soda) columns. Starting from about 30 g of rat brain, approximately 200 micrograms of pure enzyme was obtained. The procedure was very simple and highly reproducible. The enzyme thus obtained was nearly pure by silver staining after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the presence of 10% (w/v) glycerol and 0.05% (w/v) Triton X-100, the enzyme could be stored at -80 degrees C for several months.