Transgenic Research

, Volume 3, Issue 1, pp 43–49 | Cite as

Anthranilate synthase and chorismate mutase activities in transgenic tobacco plants overexpressing tryptophan decarboxylase fromCatharanthus roseus

  • Charlotte Poulsen
  • Oscar J. M. Goddijn
  • J. Harry C. Hoge
  • Robert Verpoorte
Papers

Abstract

TransgenicNicotiana tabacum L. ‘Petit Havana’ SR1 F1-plants expressing tryptophan decarboxylase cDNA (tdc) fromCatharanthus roseus (L.) G. Don under the control of the CaMV 35S promoter and terminator exhibited tryptophan decarboxylase (TDC) enzyme activity and accumulated tryptamine. The plants with the highest TDC activity contained 19 pkat per mg of protein. The influence of transgenic expression oftdc on the activities of anthranilate synthase (AS) and chorismate mutase (CM) were examined in 10 transgenic tobacco plants. The specific activities of these two chorismate-utilizing enzymes were not significantly affected by expression oftdc, despite their important functions as branch point enzymes in the shikimate pathway. The results indicate that the normal route of tryptophan biosynthesis in plants is sufficient to supply a considerable amount of this essential amino acid for the biosynthesis of secondary metabolites. Despite their increased tryptamine content, the growth and development of the transgenic tobacco plants expressingtdc appeared normal.

Keywords

tryptophan decarboxylase anthranilate synthase chorismate mutase transgenic tobacco 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Belser, W.L., Baron Murphy, J., Delmer, D.P. and Mills, S.E. (1971) End product control of tryptophan biosynthesis in extracts and intact cells of the higher plantNicotiana tabacum var. Wisconsin 38.Biochim. Biophys. Acta 237, 1–10.Google Scholar
  2. Berlin, J., Rügenhagen, C., Dietze, P., Fecker, L., Goddijn, O.J.M. and Hoge, J.H.C. (1993) Increased production of serotonin by suspension and root cultures ofPeganum harmala transformed with a tryptophan decarboxylase cDNA clone fromCatharanthus roseus. Transgenic Res. (In press).Google Scholar
  3. Brotherton, J.E., Hauptmann, R.M. and Widholm, J.M. (1986) Anthranilate synthase forms in plants and cultured cells ofNicotiana tabacum L.Planta 168, 214–21.Google Scholar
  4. Eilert, U., De Luca, V., Constabel, F. and Kurz, W.G.W. (1987) Elicitor-mediated induction of tryptophan decarboxylase and strictosidine synthase activities in cell suspension cultures ofCatharanthus roseus.Arch. Biochem. Biophys. 254, 491–7.Google Scholar
  5. Gilchrist, D.G. and Kosuge, T. (1980) Aromatic amino acid biosynthesis and its regulation. In Mifflin, B.J. ed.,The Biochemistry of Plants, Vol. 5, pp. 507–31, London: Academic Press.Google Scholar
  6. Goddijn, O.J.M. van der Duyn Schouten, P.M., Schilperoort, R.A. and Hoge, J.H.C. (1993) A chimaeric trytophan decarboxylase gene as a novel selectable marker in plant cells.Pl. Mol. Biol. (In press).Google Scholar
  7. Görisch, H. (1978) A new test for chorismate mutase activity.Anal. Biochem. 86, 764–8.Google Scholar
  8. Knobloch, K.H., Hansen, B. and Berlin, J. (1981) Mediuminduced formation of indole alkaloids and concomitant changes of interrelated enzyme activities in cell suspension cultures ofCatharanthus roseus.Z. Naturforsch. 36c, 40–3.Google Scholar
  9. Mérillon, J.M., Doireau, P., Guillot, A., Chénieux, J.C. and Rideau, M. (1986) Indole alkaloid accumulation and tryptophan decarboxylase activity inCatharanthus roseus cells cultured in three different media.Plant Cell Rep. 5, 23–6.Google Scholar
  10. Noé, W. and Berlin, J. (1985) Induction ofde-novo synthesis of tryptophan decarboxylase in cell suspensions ofCatharanthus roseus.Planta 166, 500–4.Google Scholar
  11. Pennings, E.J.M., Hegger, I., Van der Heijden, R., Duine, J.A. and Verpoorte, R. (1987) Assay of tryptophan decarboxylase fromCatharanthus roseus plant cell cultures by highperformance liquid chromatography.Anal. Biochem. 165, 133–6.Google Scholar
  12. Peterson, G.L. (1977) A simplification of the protein assay method of Lowryet al. which is more generally applicable.Anal. Biochem. 83, 346–56.Google Scholar
  13. Poulsen, C. and Verpoorte, R. (1991) Roles of chorismate mutase, isochorismate synthase and anthranilate synthase in plants.Phytochemistry 30, 377–86.Google Scholar
  14. Poulsen, C., Pennings, E.J.M. and Verpoorte, R. (1991) Highperformance liquid chromatography assay of anthranilate synthase from plant cell cultures.J. Chromatogr. 547, 155–60.Google Scholar
  15. Sasse, F., Buchholz, M., and Berlin, J. (1983) Selection of cell lines ofCatharanthus roseus with increased tryptophan decarboxylase activity.Z. Naturforsch. 38c, 916–22.Google Scholar
  16. Scott, A.I., Mizukami, H. and Lee, S.-L. (1979) Characterization of a 5-methyltryptophan resistan strain ofCatharanthus roseus cultured cells.Phytochemistry 18, 795–8.Google Scholar
  17. Songstad, D.D., De Luca, V., Brisson, N., Kurz, W.G.W. and Nessler, C.L. (1990) High levels of tryptamine accumulation in transgenic tobacco expressing tryptophan decarboxylase.Plant Physiol. 94, 1410–3.Google Scholar
  18. Widholm, J.M. (1972a) CulturedNicotiana tabacum cells with an altered anthranilate synthetase which is less sensitive to feedback inhibition.Biochim. Biophys. Acta 261, 52–8.Google Scholar
  19. Widholm, J.M. (1972b) Anthranilate synthetase from 5-methyl-tryptophan-susceptible and-resistant culturedDaucus carota cells.Biochim. Biophys. Acta 279, 48–57.Google Scholar
  20. Widholm, J.M. (1974) Control of aromatic amino acid biosynthesis in cultured plant tissues: effect of intermediates and aromatic amino acids on free levels.Physiol. Plant. 30, 13–8.Google Scholar
  21. Zalkin, H. (1980) Anthranilate synthase: relationships between bifunctional and monofunctional enzymes. In Bisswanger, H. and Schmincke-Ott, E. eds.,Multifunctional Proteins, pp. 123–49. John Wiley & Sons, New York.Google Scholar

Copyright information

© Chapman & Hall 1994

Authors and Affiliations

  • Charlotte Poulsen
    • 1
    • 2
  • Oscar J. M. Goddijn
    • 3
  • J. Harry C. Hoge
    • 3
  • Robert Verpoorte
    • 2
  1. 1.Danisco A/SCopenhagen KDenmark
  2. 2.Project Group Plant Cell Biotechnology Biotechnology Delft Leiden, Center for Bio-Pharmaceutical Sciences Division of PharmacognosyUniversity of LeidenLeidenThe Netherlands
  3. 3.Institute of Molecular Plant Sciences, Clusius LaboratoryUniversity of LeidenLeidenThe Netherlands

Personalised recommendations