Identification of two isoenzymes of protein phosphatase 2C in both rabbit skeletal muscle and liver.
Protein phosphatase 2C was isolated from rabbit skeletal muscle by a procedure that involved chromatography on DEAE-cellulose, precipitation with ammonium sulphate, gel-filtration on Sephadex G-100, affinity chromatography on thiophosphorylated myosin-P-light-chain--Sepharose and chromatography on Mono Q. The enzyme was purified about 35,000-fold and 0.3-0.4 mg was isolated from 2500 g skeletal muscle within 5 days. The final step resolved the activity into two peaks, termed protein phosphatases 2C1 and 2C2, that possessed identical substrate specificities and enzymatic properties. About 2.5-fold more protein phosphatase 2C2 was isolated than protein phosphatase 2C1. Protein phosphatases 2C1 and 2C2 migrated as single bands on SDS/polyacrylamide gels yielding apparent molecular masses of 44 kDa and 42 kDa, respectively, and the native proteins were both monomeric at pH 7.5 as judged by their elution from Sephadex G-100 and Sephacryl S200. Peptide maps of protein phosphatases 2C1 and 2C2, obtained after separate digestions with four different proteinases, were different, indicating that they are isoenzymes. Protein phosphatases 2C1 and 2C2 were purified from rabbit liver by the same procedure, and 0.2 mg (2C1 + 2C2) was isolated from 120 g hepatic tissue. Hepatic protein phosphatases 2C1 and 2C2 were also isolated in a molar ratio of about 1:2.5, and their enzymatic properties and apparent molecular masses in the presence and absence of SDS were identical to the skeletal muscle enzymes. Protein phosphatases 2C1 from muscle and liver displayed identical peptide maps, as did protein phosphatases 2C2 from these two tissues. It is concluded that the same two isoenzymes of protein phosphatase 2C are present in skeletal muscle and liver.
D000818:Animals; D002845:Chromatography / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D007527:Isoenzymes / Q000302:isolation & purification*; D008099:Liver / Q000201:enzymology*; D009132:Muscles / Q000201:enzymology*; D010447:Peptide Hydrolases; D010449:Peptide Mapping; D010749:Phosphoprotein Phosphatases / Q000302:isolation & purification*; D011817:Rabbits; D013379:Substrate Specificity
D007527:Isoenzymes; D010749:Phosphoprotein Phosphatases; D010447:Peptide Hydrolases