Isolation of Histone H1-Stimulated Phosphoprotein Phosphatase from Kidney and Skeletal Muscle

Protein phosphorylation is an important mechanism for the regulation of numerous mammalian cellular processes. The extent of phosphorylation of any given protein is determined by the relative activity of protein kinases and protein phosphatases that catalyze the incorporation or removal of a protein-bound phosphate, respectively (1). Much is known about the regulation of a number of protein kinases (1) and it is becoming increasingly clear that protein phosphatases are also important sites for the regulation of cell function (2). Protein phosphatase activity is regulated by extrinsic (substrate-directed) and intrinsic (phosphatase-directed) mechanisms (1, 3). In the former, various metabolites bind to the phosphoprotein substrate, thus altering their susceptibility to dephosphorylation. In the latter, various ligands bind to the phosphatase and thus directly alter phosphatase activity.

Studies on protein phosphatases are complicated by the presence of multiple forms which dephosphorylate a variety of phosphoprotein substrates (1–3). Cohen and co-workers (2) have classified protein phosphatases as type 1 and type 2. Type 1 phosphatase is inhibited by heat-stable inhibitor proteins, is relatively insensitive to inhibition by ATP, and preferentially dephosphorylates the β subunit of phosphorylase kinase. Type 2 phosphatase is not inhibited by heat-stable inhibitors, is sensitive to inhibition by ATP, and preferentially dephosphorylates the α subunit of phosphorylase kinase. Ingebritsen and Cohen (2) have suggested that nearly all of the protein phosphatase activity of mammalian tissues can be attributed to four multifunctional phosphatases that they have designated protein phosphatases 1 (a type 1 phosphatase) and 2A, 2B, and 2C (type 2 phosphatases). The activity of phosphatase 2C is believed to be regulated by Mg²⁺, and phosphatase 2B is dependent upon Ca²⁺ and calmodulin for activity. Phosphatase 1 appears to be the major form of protein phosphatase in skeletal muscle (4).