Domains of phosphatase inhibitor-2 involved in the control of the ATP-Mg-dependent protein phosphatase

J Biol Chem. 1994 Nov 18;269(46):28919-28.

Abstract

Inhibitor-2 (I-2) inhibits the free catalytic subunit of type 1 phosphatase (CS1) and controls the cyclic inactivation/activation of CS1 in the ATP-Mg-dependent protein phosphatase complex. We report here the effect of mutations on these two properties of I-2. Substitution of Thr-72 with Ala, Asp, or Glu generated complexes with CS1 that could not be activated. Mutation of Ser-86 did not affect activation by glycogen synthase kinase-3 (GSK-3) alone but impaired synergistic activation by casein kinase II and GSK-3. Mutations in the region between Thr-72 and Ser-86 did not alter the inhibitory potency of I-2 but prevented complete inactivation of CS1. A mutant without the 35 NH2-terminal residues exhibited an IC50 for CS1 200-fold higher than that of wild-type I-2. However, it formed an inactive phosphatase complex with CS1, which was activated by GSK-3. A mutant with the 59 COOH-terminal residues deleted retained full inhibitory activity and formed an inactive complex that could not be activated by GSK-3. We conclude that the NH2-terminal region of I-2 is involved in inhibition, that the sequence between Thr-72 and Ser-86 is necessary for the conversion of CS1 from an active to an inactive conformation, and that the COOH terminus is required for activation by GSK-3. Thus, different functional domains of I-2 may interact with distinct regions of CS1.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenosine Triphosphate / metabolism*
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Binding Sites
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism
  • Casein Kinase II
  • Glycogen Synthase Kinase 3
  • Glycogen Synthase Kinases
  • Molecular Sequence Data
  • Mutation
  • Oligodeoxyribonucleotides
  • Phosphoprotein Phosphatases / antagonists & inhibitors
  • Phosphoprotein Phosphatases / genetics
  • Phosphoprotein Phosphatases / metabolism*
  • Phosphorylation
  • Proline / genetics
  • Protein-Serine-Threonine Kinases / metabolism
  • Proteins / metabolism*
  • Rabbits
  • Sequence Deletion

Substances

  • Oligodeoxyribonucleotides
  • Proteins
  • protein phosphatase inhibitor-2
  • Adenosine Triphosphate
  • Proline
  • Glycogen Synthase Kinases
  • Casein Kinase II
  • Protein-Serine-Threonine Kinases
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Glycogen Synthase Kinase 3
  • Phosphoprotein Phosphatases