Purification and Characterization of Pyruvate‐Dehydrogenase Phosphatase from Pig‐Heart Muscle

Abstract

Pyruvate‐dehydrogenase phosphatase was extensively purified from pig heart muscle. The molecular weight was determined to be in the range between 92000 and 95000. By dodecylsulfate‐gel electrophoresis there was no indication for a subunit structure of the protein. The phosphatase is dependent on Mg 2+ or Mn 2+ . Half‐maximal activity was obtained, in histidine buffer, at 2.5 mM MgCl 2 and 1.8 mM MnCl 2 , respectively. Fluoride at a concentration of 0.6 mM inhibited phosphatase activity by 50% in the presence of Mg 2+ , but not of Mn 2+ . The enzyme was not affected by alkylating thiol‐reagents. Studies with sequestering agents suggest that pyruvate‐dehydrogenase phosphatase contains a metal which, besides Mg 2+ or Mn 2+ , is essential for its function.