Tyrosine-272 is involved in the inhibition of protein phosphatase-1 by multiple toxins

Biochemistry. 1996 Feb 6;35(5):1606-11. doi: 10.1021/bi9521396.

Abstract

Protein phosphatase-1 (PP1) is regulated by interaction with different subunits, which include several inhibitory proteins. It is also potently inhibited by several toxins of diverse origins. Recent work has identified a region near the C-terminus of PP1 (residues 274-277) whose modification was shown to moderate okadaic acid binding [Zhang et al. (1994) J. Biol. Chem. 269, 16997-17000]. In this study, the role of this region in toxin binding was explored by site-directed mutagenesis. A residue (Tyr-272) was identified whose mutation had dramatic effects on the spectrum of inhibitor sensitivity of PP1. The IC50's of a number of mutants of Tyr-272 toward okadaic acid, tautomycin, calyculin A, microcystin-LR, nodularin, inhibitor-2, and cantharidic acid were determined and compared to that of the wild-type enzyme. The sensitivity of PP1 toward tautomycin and calyculin A was markedly decreased, by as much as 3 orders of magnitude, with lesser effects on okadaic acid and nodularin, and with microcystin-LR and inhibitor-2 being the least affected. These studies show that Tyr-272 is of specific importance for the binding of these inhibitors and provide strong evidence for the postulate that these toxins all bind to a common inhibitor site on PP1. In addition, our studies show that Tyr-272 is not required for catalytic activity.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Cysteine
  • DNA Mutational Analysis
  • Drug Resistance
  • Enzyme Inhibitors / pharmacology*
  • Ethers, Cyclic / pharmacology
  • Molecular Sequence Data
  • Okadaic Acid
  • Phosphoprotein Phosphatases / antagonists & inhibitors*
  • Phosphoprotein Phosphatases / genetics
  • Protein Binding / genetics
  • Protein Phosphatase 1
  • Structure-Activity Relationship
  • Toxins, Biological / pharmacology*
  • Tyrosine

Substances

  • Enzyme Inhibitors
  • Ethers, Cyclic
  • Toxins, Biological
  • Okadaic Acid
  • Tyrosine
  • Phosphoprotein Phosphatases
  • Protein Phosphatase 1
  • Cysteine