Purification and Subunit Structure of a High-Molecular-Weight Phosphoprotein Phosphatase (Phosphatase 11) from Rat Liver

Abstract

1 Phosphatase II is a form of phosphoprotein phosphatase originally found in rat liver extract; it has a molecular weight of 160000 by gel filtration and is highly active towards phosphorylase a. This phosphatase has been purified 1800‐fold by using DEAE‐cellulose (DE‐52), aminohexyl–Sepharose‐4B, protamine–Sepharose4B and Sephadex G‐200 chromatography. Throughout the purification steps, the original molecular weight and substrate specificity of phosphatase II were almost perfectly preserved. 2 The product of the final purification step migrated predominantly as a single protein band on non‐denaturing gel electrophoresis. Sodium dodecyl sulfate gel electorphoresis revealed that the enzyme contains two types of subunit, α and β, with molecular weights of 35000 and 69000, respectively. When treated with 0.2 M 2‐mercaptoethanol at −20°C, phosphatase II was dissociated to release the catalytically active α subunit. The β subunit may be catalytically inactive but interacts with the α subunit so that phosphatase I1 becomes much less susceptible than the α subunit to inactivation by ATP or pyrophosphate.