Glutathionylation of human thioredoxin: A possible crosstalk between the glutathione and thioredoxin systems

Simona Casagrande, Valentina Bonetto, Maddalena Fratelli, Elisabetta Gianazza, Ivano Eberini, Tania Massignan, Mario Salmona, Geng Chang, Arne Holmgren, Pietro Ghezzi

Research output: Contribution to journalArticlepeer-review


To identify proteins undergoing glutathionylation (formation of protein-glutathione mixed disulfides) in human T cell blasts, we radiolabeled the glutathione pool with 35S, exposed cells to the oxidant diamide, and analyzed cellular proteins by two-dimensional electrophoresis. One of the proteins undergoing glutathionylation was identified by molecular weight, isoelectric point, and immunoblotting as thioredoxin (Trx). Incubation of recombinant human Trx with glutathione disulfide or S-nitrosoglutathione led to the formation of glutathionylated Trx, identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry. The glutathionylation site was identified as Cys-72. Glutathionylation of rhTrx abolished its enzymatic activity as insulin disulfide reductase in the presence of NADPH and Trx reductase. Activity was, however, regained with sigmoidal kinetics, indicating a process of autoactivation due to the ability of Trx to deglutathionylate itself. These data suggest that the intracellular glutathione/glutathione disulfide ratio, an indicator of the redox state of the cell, can regulate Trx functions reversibly through thiol-disulfide exchange reactions.

Original languageEnglish
Pages (from-to)9745-9749
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number15
Publication statusPublished - Jul 23 2002

ASJC Scopus subject areas

  • Genetics
  • General


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