An atypical catalytic mechanism involving three cysteines of thioredoxin

Abstract

Unlike other thioredoxins h characterized so far, a poplar thioredoxin of the h type, PtTrxh4, is reduced by glutathione and glutaredoxin (Grx) but not NADPH:thioredoxin reductase (NTR). PtTrxh4 contains three cysteines: one localized in an N-terminal extension (Cys4) and two (Cys58 and Cys61) in the classical thioredoxin active site ( 57WCGPC61). The property of a mutant in which Cys 58 was replaced by serine demonstrates that it is responsible for the initial nucleophilic attack during the catalytic cycle. The observation that the C4S mutant is inactive in the presence of Grx but fully active when dithiothreitol is used as a reductant indicates that Cys4 is required for the regeneration of PtTrxh4 by Grx. Biochemical and x-ray crystallographic studies indicate that two intramolecular disulfide bonds involving Cys 58 can be formed, linking it to either Cys61 or Cys 4. We propose thus a four-step disulfide cascade mechanism involving the transient glutathionylation of Cys4 to convert this atypical thioredoxin h back to its active reduced form. © 2008 by The American Society for Biochemistry and Molecular Biology, Inc.

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Citation

Cha, S.K., Navrot, N., Didierjean, C., Rouhier, N., Hirasawa, M., Knaff, D.B., Wingsle, G., Samian, R., Jacquot, J.-P., Corbier, C., & Gelhaye, E.. 2008. An atypical catalytic mechanism involving three cysteines of thioredoxin. Journal of Biological Chemistry, 283(34). https://doi.org/10.1074/jbc.M802093200

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