Biochemical and molecular characterization of the mitochondrial peroxiredoxin PsPrxII F from Pisum sativum

Plant Physiol Biochem. Oct-Nov 2007;45(10-11):729-39. doi: 10.1016/j.plaphy.2007.07.017. Epub 2007 Aug 7.

Abstract

The pea peroxiredoxin homologue PsPrxII F of the Arabidopsis thaliana mitochondrial AtPrxII F was isolated as cDNA and genomic DNA, and characterized in respect to its biochemical and molecular properties. The deduced amino acid sequence contains an N-terminal targeting address for mitochondrial import. Mitochondrial location of PsPrxII F was confirmed by immunocytochemistry. The mature enzyme, without the transit peptide, has a molecular mass of 18.75 kDa, and, at positions 59 and 84, carries the two catalytic cysteinyl residues which are characteristic for this particular Prx subgroup. Activity of site-directed mutagenized C84S-variant lacking the so-called resolving Cys dropped to about 12% of WT Prx while C59S lost its peroxidatic activity completely. Likewise, WT PsPrxII F and C84S-variant but not C59S protected plasmid DNA against strand breakage in a mixed function oxidation assay. WT PrxII F and the variant proteins aggregated to high mass oligomers not yet described for type II Prx. Upon oxidation with hydrogen peroxide PsPrxII F focussed in a series of spots of distinct pI but similar molecular masses in two-dimensional gels indicating different oxidation states of the protein. Using this technique, partial oxidation was also detected in leaf extracts and isolated mitochondria. PsPrxII F mRNA and protein accumulated in cold and heavy metals treated pea plants suggesting a particular function under stress.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Blotting, Western
  • Cold Temperature
  • Electrophoresis, Gel, Two-Dimensional
  • Hydrogen Peroxide / pharmacology
  • Immunohistochemistry
  • Metals, Heavy / pharmacology
  • Mitochondrial Proteins / genetics
  • Mitochondrial Proteins / metabolism*
  • Models, Genetic
  • Molecular Sequence Data
  • Oxidation-Reduction / drug effects
  • Oxidative Stress / drug effects
  • Peas / drug effects
  • Peas / genetics
  • Peas / metabolism*
  • Peroxiredoxins / genetics
  • Peroxiredoxins / metabolism*
  • Plant Proteins / genetics
  • Plant Proteins / metabolism*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Signal Transduction / drug effects

Substances

  • Metals, Heavy
  • Mitochondrial Proteins
  • Plant Proteins
  • RNA, Messenger
  • Hydrogen Peroxide
  • Peroxiredoxins

Associated data

  • GENBANK/AM411615