CYP79F1 and CYP79F2 have distinct functions in the biosynthesis of aliphatic glucosinolates in Arabidopsis

Authors

  • Sixue Chen,

    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
    • Present address: Department of Biology, Plant Science Institute, University of Pennsylvania, 3800 Hamilton Walk, Philadelphia, PA 19104, USA.

  • Erich Glawischnig,

    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
  • Kirsten Jørgensen,

    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
  • Peter Naur,

    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
  • Bodil Jørgensen,

    1. The Biotechnology Group, Danish Institute of Agricultural Science, The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
  • Carl-Erik Olsen,

    1. Chemistry Department, The Royal Veterinary and Agricultural University, Thorvaldsensvej 40, DK-1871 Frederiksberg C, Copenhagen, Denmark, and
    Search for more papers by this author
  • Carsten H. Hansen,

    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    Search for more papers by this author
  • Hasse Rasmussen,

    1. IACR-Rothamsted, Harpenden AL5 2JQ, UK
    Search for more papers by this author
  • John A. Pickett,

    1. IACR-Rothamsted, Harpenden AL5 2JQ, UK
    Search for more papers by this author
  • Barbara A. Halkier

    Corresponding author
    1. Plant Biochemistry Laboratory, Department of Plant Biology, and Center for Molecular Plant Physiology (PlaCe), The Royal Veterinary and Agricultural University, Copenhagen, Denmark,
    • For correspondence (fax +45 35 28 33 33; e-mail bah@kvl.dk).

    Search for more papers by this author

Summary

Cytochromes P450 of the CYP79 family catalyze the conversion of amino acids to oximes in the biosynthesis of glucosinolates, a group of natural plant products known to be involved in plant defense and as a source of flavor compounds, cancer-preventing agents and bioherbicides. We report a detailed biochemical analysis of the substrate specificity and kinetics of CYP79F1 and CYP79F2, two cytochromes P450 involved in the biosynthesis of aliphatic glucosinolates in Arabidopsis thaliana. Using recombinant CYP79F1 and CYP79F2 expressed in Escherichia coli and Saccharomyces cerevisiae, respectively, we show that CYP79F1 metabolizes mono- to hexahomomethionine, resulting in both short- and long-chain aliphatic glucosinolates. In contrast, CYP79F2 exclusively metabolizes long-chain elongated penta- and hexahomomethionines. CYP79F1 and CYP79F2 are spatially and developmentally regulated, with different gene expression patterns. CYP79F2 is highly expressed in hypocotyl and roots, whereas CYP79F1 is strongly expressed in cotyledons, rosette leaves, stems, and siliques. A transposon-tagged CYP79F1 knockout mutant completely lacks short-chain aliphatic glucosinolates, but has an increased level of long-chain aliphatic glucosinolates, especially in leaves and seeds. The level of long-chain aliphatic glucosinolates in a transposon-tagged CYP79F2 knockout mutant is substantially reduced, whereas the level of short-chain aliphatic glucosinolates is not affected. Biochemical characterization of CYP79F1 and CYP79F2, and gene expression analysis, combined with glucosinolate profiling of knockout mutants demonstrate the functional role of these enzymes. This provides valuable insights into the metabolic network leading to the biosynthesis of aliphatic glucosinolates, and into metabolic engineering of altered aliphatic glucosinolate profiles to improve nutritional value and pest resistance.

Ancillary