The role of ATP and divalent cations in the regulation of a cardiac phosphorylase phosphatase (phosphoprotein phosphatase) of Mr = 35,000.
The effects of ATP and divalent cations on a divalent cation-independent phosphorylase phosphatase of Mr = 35,000 (phosphatase S) purified from canine cardiac muscle have been studied. The enzyme can be rapidly inactivated by ATP or other nucleoside di- and triphosphates and PPi, but not by AMP, adenosine, adenine, Pi, EDTA, ethylene glycol bis(beta-aminoethyl ether)N,N' -tetraacetic acid, 1,10-phenanthroline, or 8-hydroxyquinoline. After removing the inactivating agent, such as ATP or PPi, by gel filtraiton followed by exhaustive dialysis, the inactivated enzyme (apophosphatase S) can be reactivated by preincubating with Mn2+ or Co2+, but not with Mg2+, Ca2+, Ni2+, Zn2+, Fe2+, Cu2+, Ba2+, Hg2+, Pb2+, or Cd2+. The Mn2+ -reactivated enzyme, which is less active than the Co2+ -reactivated enzyme, can be again inactivated by preincubating with ATP. The present findings indicate that phosphatase S contains a tightly bound divalent cation, probably Mn2+, in the active site. ATP and PPi, due to their structural similarity to the phosphoprotein substrate and their ability to chelate metal ions, can readily enter the active site to remove the divalent cation(s) essential for the catalytic function. The present findings also indicate that phosphatase S, a common catalytic subunit of several larger molecular forms of nospecific phosphoprotein phosphatase in cardiac muscle, can exist in two interconvertible forms, a metallized form (active) and a demetallized form (inactive). ATP and metal ions may regulate this class of isozymes by mediating the interconversions.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002413:Cations, Divalent; D003035:Cobalt / Q000494:pharmacology; D004285:Dogs; D004355:Drug Stability; D004789:Enzyme Activation; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D009206:Myocardium / Q000201:enzymology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010765:Phosphorylase Phosphatase / Q000378:metabolism*; D013329:Structure-Activity Relationship
D002413:Cations, Divalent; D003035:Cobalt; D008345:Manganese; D000255:Adenosine Triphosphate; D010749:Phosphoprotein Phosphatases; D010765:Phosphorylase Phosphatase