A phosphoprotein phosphatase from ox brain

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Abstract
A phosphoprotein phosphatase acting on a-casein has been purified from ox brain 100-fold and in a yield of 25%. The pH optimum of the enzyme is 5.5. It is activated by Mg2+ or Mn2+ ions but not by thiol compounds. In the absence of Mg2+ ions Km = 37 mg atoms of [alpha]-casein phosphorus/1. The enzyme lost activity when dialysed against H2O salt solutions. With crude preparations the loss was prevented by dialysis against cysteine (20 mM). With more purified preparations cysteine did not prevent the loss of activity on dialysis. Activity was also lost during fractionation with (NH4)2SO4 and with graded concentrations of acetone. At pH 5.9 the enzyme was inactive towards aliphatic and aromatic phosphomonoesters, phospho-amides, a phosphodiester, inorganic polymetaphosphates, pyrophos-phates, polyseryl phosphates and mixed phospholipids. It was active towards [alpha]- and [beta]-casein, phosvitin, ovalbumin and preparations of brain phosphoprotein but not towards hexokinase or pepsin. Phospho-peptones prepared from [alpha]-casein were attaked by the enzyme. The enzyme liberated only a small proportion of the total phosphate from [alpha]-casein and phosvitin before action ceased. The phosphorus released from phosvitin included 85% of the radioactive phosphorus incorporated by incubation with a protein phosphokinase. The brain enzyme differs from phosphoprotein phosphatases derived from spleen or liver.