ATP x Mg-dependent protein phosphatase from rabbit skeletal muscle. II. Purification of the activating factor and its characterization as a bifunctional protein also displaying synthase kinase activity.
A protein (FA) has been isolated from rabbit muscle which has two functions: one is the activation of the ATP x Mg-dependent phosphatase (see previous paper) (1) and the second is the phosphorylation and concomitant inactivation of glycogen synthase, independent from cyclic AMP or Ca ions. The two activities co-purify throughout the purification scheme, and reside in the single protein band that the purified preparation shows in discontinuous acrylamide gel electrophoresis. Heat inactivation experiments with the purified protein showed a parallel decrease of both activities with time. GTP could efficiently replace the ATP in both reactions. Sodium dodecyl sulfate-gel electrophoresis also shows a single protein-stained band corresponding to a Mr = approximately 50,000 and sucrose density gradient centrifugation gave a value of 45,000. The enzyme incorporates only 1 mol of phosphate/mol of synthase monomer (85,000 daltons) and brings the activity ratio (+/- glucose-6-P) down to less than 0.05. Kinetic studies suggest that FA exerts its two activities in quite different ways: the activation of the ATP x Mg-dependent phosphatase is bought about by a protein-protein interaction (FA x FC complex formation) with ATP x Mg as a necessary cofactor, whereas for the inactivation of synthase, FA is a cyclic AMP- and Ca-independent kinase.
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D000242:Cyclic AMP / Q000494:pharmacology; D004789:Enzyme Activation; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010764:Phosphorylase Kinase / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D011817:Rabbits
D011506:Proteins; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D010764:Phosphorylase Kinase; D010749:Phosphoprotein Phosphatases; D008274:Magnesium