Characterization of plant sulfiredoxin and role of sulphinic form of 2-Cys peroxiredoxin

J Exp Bot. 2010 Mar;61(5):1509-21. doi: 10.1093/jxb/erq016. Epub 2010 Feb 22.

Abstract

The antioxidant function of 2-Cys peroxiredoxin (Prx) involves the oxidation of its conserved peroxidatic cysteine to sulphenic acid that is recycled by a reductor agent. In conditions of oxidative stress, the peroxidatic cysteine can be overoxidized to sulphinic acid inactivating the Prx. An enzyme recently discovered, named sulfiredoxin (Srx), reduces the sulphinic 2-Cys Prx (Prx-SO(2)H). To explore the physiological functions of Srx in plants we have cloned, expressed and purified to homogeneity a Srx from Arabidopsis thaliana (AtSrx), as well as five variants by site-directed mutagenesis on amino acids involved in its activity. The activity of sulfiredoxin, determined by a new method, is dependent on the concentration of the sulphinic form of Prx and the conserved Srx is capable of regenerating the functionality of both pea and Arabidopsis Prx-SO(2)H. Molecular modelling of AtSrx and the facts that the R28Q variant shows a partial inactivation, that the activity of the E76A variant is equivalent to that of the native enzyme and that the double mutation R28Q/E76A abolishes the enzymatic activity suggests that the pair His100-Glu76 may be involved in the activation of C72 in the absence of R28. The knock-out mutant plants without Srx or 2-Cys Prx exhibited phenotypical differences under growth conditions of 16 h light, probably due to the signalling role of the sulphinic form of Prx. These mutants showed more susceptibility to oxidative stress than wild-type plants. This work presents the first systematic biochemical characterization of the Srx/Prx system from plants and contributes to a better understanding of its physiological function.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Arabidopsis / genetics
  • Arabidopsis / metabolism*
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism*
  • Blotting, Western
  • Electrophoresis, Polyacrylamide Gel
  • Hydrogen Peroxide / metabolism
  • Kinetics
  • Mutagenesis, Site-Directed
  • Oxidoreductases Acting on Sulfur Group Donors / genetics
  • Oxidoreductases Acting on Sulfur Group Donors / metabolism*
  • Peas / genetics
  • Peas / metabolism
  • Peroxiredoxins / genetics
  • Peroxiredoxins / metabolism*
  • Plants, Genetically Modified / genetics
  • Plants, Genetically Modified / metabolism
  • Polymerase Chain Reaction

Substances

  • Arabidopsis Proteins
  • Hydrogen Peroxide
  • Peroxiredoxins
  • Oxidoreductases Acting on Sulfur Group Donors
  • sulfiredoxin protein, Arabidopsis