Partial purification and identification of GDP-mannose 3",5"-epimerase of Arabidopsis thaliana, a key enzyme of the plant vitamin C pathway

Proc Natl Acad Sci U S A. 2001 Dec 18;98(26):14843-8. doi: 10.1073/pnas.011578198.


The first step in the biosynthetic pathway of vitamin C in plants is the formation, at the level of sugar nucleotide, of l-galactosyl residues, catalyzed by a largely unknown GDP-d-mannose 3",5"-epimerase. By using combined conventional biochemical and mass spectrometry methods, we obtained a highly purified preparation of GDP-d-mannose 3",5"-epimerase from an Arabidopsis thaliana cell suspension. The native enzyme is an 84-kDa dimer, composed of two apparently identical subunits. In-gel tryptic digestion of the enzyme subunit, followed by peptide sequencing and a blast search, led to the identification of the epimerase gene. The closest homolog of the plant epimerase is the BlmG gene product of Streptomyces sp., a putative NDP-d-mannose 5"-epimerase. The plant GDP-d-mannose 3",5"-epimerase is, to our knowledge, a novel member of the extended short-chain dehydrogenase/reductase family. The enzyme was cloned and expressed in Escherichia coli cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / enzymology*
  • Arabidopsis / metabolism
  • Ascorbic Acid / metabolism*
  • Base Sequence
  • Carbohydrate Epimerases / chemistry
  • Carbohydrate Epimerases / isolation & purification*
  • Carbohydrate Epimerases / metabolism
  • Chromatography, Ion Exchange
  • DNA Primers
  • Molecular Sequence Data
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Spectrometry, Mass, Electrospray Ionization
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization


  • DNA Primers
  • Recombinant Proteins
  • Carbohydrate Epimerases
  • GDPmannose 3,5-epimerase
  • Ascorbic Acid