Purification and properties of phosphoprotein phosphatase from rabbit heart

Abstract

A low molecular weight phosphoprotein phosphatase acting on muscle phosphorylase a has been purified to homogeneity from rabbit heart by acid precipitation, ethanol treatment, and chromatography on Sephadex G-75 and Sepharose-histone. The purified enzyme showed a single band when examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis; the molecular weight calculated by this method was 34,000. The S20,w value and Stockes radius for the enzyme were 3.45 and 24.0 A, respectively. Using these two values, a molecular weight of 35,000 was calculated. Purified enzyme showed a wide substrate specificity and catalyzed the dephosphorylation of phosphorylase a, glycogen synthase D, phosphorylated histone, and phosphorylated casein. A heat-stable protein inhibitor for this enzyme with phosphorylase a as the substrate was also shown to be present in crude extracts of rabbit heart. It inhibited the dephosphorylation of phosphorylase a by phosphoprotein phosphatase by decreasing the Vmax of the reaction.