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Neurochemical Research

, Volume 36, Issue 2, pp 232–240 | Cite as

Phenylarsine Oxide Binding Reveals Redox-Active and Potential Regulatory Vicinal Thiols on the Catalytic Subunit of Protein Phosphatase 2A

  • Timothy D. FoleyEmail author
  • Scott L. Melideo
  • Adriana E. Healey
  • Eugene J. Lucas
  • Jason A. Koval
ORIGINAL PAPER

Abstract

Our earlier finding that the activity of protein phosphatase 2A from rat brain is inhibited by micromolar concentrations of the dithiol cross-linking reagent phenylarsine oxide (PAO) has encouraged the hypothesis that the catalytic subunit (PP2Ac) of PP2A contains one or more pairs of closely-spaced (vicinal) thiol pairs that may contribute to regulation of the enzyme. The results of the present study demonstrate using immobilized PAO-affinity chromatography that PP2Ac from rat brain formed stable DTT-sensitive adducts with PAO with or without associated regulatory subunits. In addition, a subset of the PAO-binding vicinal thiols of PP2Ac was readily oxidized to disulfide bonds in vitro. Importantly, a small fraction of PP2Ac was still found to contain disulfide bonds after applying stringent conditions designed to prevent protein disulfide bond formation during homogenization and fractionation of the brains. These findings establish the presence of potentially regulatory and redox-active PAO-binding vicinal thiols on the catalytic subunit of PP2A and suggest that a population of PP2Ac may contain disulfide bonds in vivo.

Keywords

Protein phosphatase 2A Disulfide bonds Phenylarsine oxide Oxidative stress 

Abbreviations

DTT

Dithiothreitol

EtOH-AS65

The ethanol and ammonium sulfate-precipitated fraction containing dissociated PP2Ac

PAO

Phenylarsine oxide

PP2A

Protein phosphatase 2A

PP2Ac

Protein phosphatase 2A catalytic subunit

IAM

Iodoacetamide

S100

The 100,000g supernatant

TCEP

Tris(2-carboxyethyl)-phosphine

Notes

Acknowledgments

This work was supported in part by NIH Grant AG022357 from the National Institute on Aging.

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Timothy D. Foley
    • 1
    Email author
  • Scott L. Melideo
    • 1
  • Adriana E. Healey
    • 1
  • Eugene J. Lucas
    • 1
  • Jason A. Koval
    • 1
  1. 1.Department of Chemistry and Program in Biochemistry, Cell, and Molecular BiologyUniversity of ScrantonScrantonUSA

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