Oxidative modifications of glyceraldehyde-3-phosphate dehydrogenase play a key role in its multiple cellular functions

Na Rae Hwang, Seung Hee Yim, Young Mee Kim, Jaeho Jeong, Eun Joo Song, Yoonji Lee, Jin Hee Lee, Sun Choi, Kong Joo Lee

Research output: Contribution to journalArticle

93 Citations (Scopus)

Abstract

Knowledge of the cellular targets of ROS (reactive oxygen species) and their regulation is an essential prerequisite for understanding ROS-mediated signalling. GAPDH (glyceraldehyde-3-phosphate dehydrogenase) is known as a major target protein in oxidative stresses and becomes thiolated in its active site. However, the molecular and functional changes of oxidized GAPDH, the inactive form, have not yet been characterized. To examine the modifications of GAPDH under oxidative stress, we separated the oxidation products by two-dimensional gel electrophoresis and identified them using nanoLC-ESI-q-TOF MS/MS (nano column liquid chromatography coupled to electrospray ionization quadrupole time-of-flight tandem MS). Intracellular GAPDH subjected to oxidative stress separated into multiple acidic spots on two-dimensional gel electrophoresis and were identified as cysteine disulfide and cysteic acids on Cys152 in the active site. We identified the interacting proteins of oxidized inactive GAPDH as p54nrb (54 kDa nuclear RNA-binding protein) and PSF (polypyrimidine tract-binding protein-associated splicing factor), both of which are known to exist as heterodimers and bind to RNA and DNA. Interaction between oxidized GAPDH and p54nrb was abolished upon expression of the GAPDH active site mutant C152S. The C-terminal of p54nrb binds to GAPDH in the cytosol in amanner dependent on the dose of hydrogen peroxide. The GAPDH-p54nrb complex enhances the intrinsic topoisomerase I activation by p54nrb-PSF binding. These results suggest that GAPDH exerts other functions beyond glycolysis, and that oxidatively modified GAPDH regulates its cellular functions by changing its interacting proteins, i.e. the RNA splicing by interacting with the p54nrb-PSF complex.

Original languageEnglish (US)
Pages (from-to)253-264
Number of pages12
JournalBiochemical Journal
Volume423
Issue number2
DOIs
StatePublished - Oct 15 2009

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Glyceraldehyde-3-Phosphate Dehydrogenases
Oxidative stress
Catalytic Domain
Oxidative Stress
Electrophoresis, Gel, Two-Dimensional
Electrophoresis
Reactive Oxygen Species
Polypyrimidine Tract-Binding Protein
Cysteic Acid
Gels
RNA
RNA Splicing
Nuclear RNA
Electrospray ionization
Type I DNA Topoisomerase
Column chromatography
Proteins
RNA-Binding Proteins
Liquid chromatography
Glycolysis

Keywords

  • 54 kDa nuclear RNA-binding protein (p54nrb)
  • Active site cysteine
  • Cysteic acid
  • Glyceraldehyde-3-phosphate dehydrogenase (GAPDH)
  • Oxidative stress
  • Topoisomerase

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Hwang, N. R., Yim, S. H., Kim, Y. M., Jeong, J., Song, E. J., Lee, Y., ... Lee, K. J. (2009). Oxidative modifications of glyceraldehyde-3-phosphate dehydrogenase play a key role in its multiple cellular functions. Biochemical Journal, 423(2), 253-264. https://doi.org/10.1042/BJ20090854

Oxidative modifications of glyceraldehyde-3-phosphate dehydrogenase play a key role in its multiple cellular functions. / Hwang, Na Rae; Yim, Seung Hee; Kim, Young Mee; Jeong, Jaeho; Song, Eun Joo; Lee, Yoonji; Lee, Jin Hee; Choi, Sun; Lee, Kong Joo.

In: Biochemical Journal, Vol. 423, No. 2, 15.10.2009, p. 253-264.

Research output: Contribution to journalArticle

Hwang, NR, Yim, SH, Kim, YM, Jeong, J, Song, EJ, Lee, Y, Lee, JH, Choi, S & Lee, KJ 2009, 'Oxidative modifications of glyceraldehyde-3-phosphate dehydrogenase play a key role in its multiple cellular functions', Biochemical Journal, vol. 423, no. 2, pp. 253-264. https://doi.org/10.1042/BJ20090854
Hwang, Na Rae ; Yim, Seung Hee ; Kim, Young Mee ; Jeong, Jaeho ; Song, Eun Joo ; Lee, Yoonji ; Lee, Jin Hee ; Choi, Sun ; Lee, Kong Joo. / Oxidative modifications of glyceraldehyde-3-phosphate dehydrogenase play a key role in its multiple cellular functions. In: Biochemical Journal. 2009 ; Vol. 423, No. 2. pp. 253-264.
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