Plant Molecular Biology

, Volume 36, Issue 1, pp 75–87 | Cite as

Purification, regulation and cloning of a glutathione transferase (GST) from maize resembling the auxin-inducible type-III GSTs

  • David P. Dixon
  • David J. Cole
  • Robert Edwards


The glutathione transferases (GSTs) from maize (Zea mays L.) with activities toward the chloroacetanilide herbicide metolachlor and the diphenyl ether herbicide fluorodifen were fractionated into two pools based on binding to affinity columns. Pool 1 GSTs were retained on Orange A agarose and were identified as isoenzymes Zea mays (Zm) GST I-I, Zm GST I-II and Zm GST I-III, which have been described previously. Pool 2 GSTs selectively bound to S-hexyl-glutathione-Sepharose and were distinct from the pool 1 GSTs, being composed of a homodimer of 28.5 kDa subunits, termed Zm GST V-V, and a heterodimer of the 28.5 kDa polypeptide and a 27.5 kDa subunit, termed Zm GST V-VI. Using an antibody raised to Zm GST V-VI, a cDNA expression library was screened and a Zm GST V clone identified showing sequence similarity to the type-III auxin-inducible GSTs previously identified in tobacco and other dicotyledenous species. Recombinant Zm GST V-V showed high GST activity towards the diphenyl ether herbicide fluorodifen, detoxified toxic alkenal derivatives and reduced organic hydroperoxides. Antibodies raised to Zm GST I-II and Zm GST V-VI were used to monitor the expression of GST subunits in maize seedlings. Over a 24 h period the Zm GST I subunit was unresponsive to chemical treatment, while expression of Zm GST II was enhanced by auxins, herbicides, the herbicide safener dichlormid and glutathione. The Zm GST V subunit was more selective in its induction, only accumulating significantly in response to dichlormid treatment. During development Zm GST I and Zm GST V were expressed more in roots than in shoots, with Zm GST II expression limited to the roots.

Zea mays glutathione transferase herbicide herbicide safener 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Bollag DM, Edelstein SJ: Protein Methods. Wiley-Liss, New York (1991).Google Scholar
  2. 2.
    Cleveland DW, Fischer SG, Kirschner MW, Laemmli UK: Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem 252: 1102–1106 (1977).PubMedGoogle Scholar
  3. 3.
    Dean JV, Devarenne TP: Peroxidase-mediated conjugation of glutathione to unsaturated phenylpropanoids. Evidence against glutathione S-transferase involvement. Physiol Plant 99: 271–278 (1997).Google Scholar
  4. 4.
    Dean JV, Devarenne TP, Lee I-S, Orlofsky LE: Properties of a maize glutathione S-transferase that conjugates coumaric acid and other phenylpropanoids. Plant Physiol 108: 985–994 (1995).PubMedGoogle Scholar
  5. 5.
    Dixon D, Cole DJ, Edwards R: Characterisation of multiple glutathione transferases containing the GST I subunit with activities toward herbicide substrates in maize (Zea mays). Pestic Sci 50: 72–82 (1997).Google Scholar
  6. 6.
    Droog F: Plant glutathione S-transferases, a tale of theta and tau. J Plant Growth Regul 16: 95–107 (1997).Google Scholar
  7. 7.
    Droog FNJ, Hooykaas PJJ, Libbenga KR, van der Zaal, EJ: Proteins encoded by an auxin-regulated gene family of tobacco share limited but significant homology with glutathione Stransferases and one member indeed shows in vitro GST activity. Plant Mol Biol 21: 965–972 (1993).CrossRefPubMedGoogle Scholar
  8. 8.
    Droog FNJ, Hooykaas PJJ, van der Zaal BJ: 2,4-Dichlorophenoxyacetic acid and related chlorinated compounds inhibit two auxin-regulated type-III tobacco glutathione S-transferases. Plant Physiol 107: 1139–1146 (1995).PubMedGoogle Scholar
  9. 9.
    Edwards R: Characterization of glutathione transferases and glutathione peroxidases in pea (Pisum sativum). Physiol Plant 98: 594–604 (1996).Google Scholar
  10. 10.
    Farago S, Brunold C, Kreuz K: Herbicide safeners and glutathione metabolism. Physiol Plant 91: 537–542 (1994).Google Scholar
  11. 11.
    Fuerst EP, Irzyk GP, Miller KD: Partial characterization of glutathione S-transferase isozymes induced by the herbicide safener benoxacor in maize. Plant Physiol 102: 795–802 (1993).PubMedGoogle Scholar
  12. 12.
    Graff G, Anderson LA, Jaques LW: Preparation and purification of soybean lipoxygenase-derived unsaturated hydroperoxy and hydroxy fatty acids and determination of molar absorptivities of hydroxy fatty acids. Anal Biochem 188: 38–47 (1990).Google Scholar
  13. 13.
    Guddewar MB, Dauterman WC: Purification and properties of a glutathione S-transferase from corn which conjugates striazine herbicides. Phytochemistry 18: 735–740 (1979).Google Scholar
  14. 14.
    Harlow E, Lane D: Antibodies: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1988).Google Scholar
  15. 15.
    Hatton PJ, Dixon D, Cole DJ, Edwards R: Glutathione transferase activities and herbicide selectivity in maize and associated weed species. Pestic Sci 46: 267–275 (1996).Google Scholar
  16. 16.
    Holt DC, Lay VJ, Clarke ED, Dinsmore A, Jepson, I, Bright SWJ, Greenland AJ: Characterization of the safener-induced glutathione S-transferase isoform II from maize. Planta 196: 295–302 (1995).PubMedGoogle Scholar
  17. 17.
    Irzyk G, Potter S, Ward E, Fuerst EP: A cDNA clone encoding the 27-kilodalton subunits of glutathione S-transferase IV from Zea mays. Plant Physiol 107: 311–312 (1995).PubMedGoogle Scholar
  18. 18.
    Irzyk GP, Fuerst EP: Purification and characterization of a glutathione S-transferase from benoxacor-treated maize (Zea mays). Plant Physiol 102: 803–810 (1993).PubMedGoogle Scholar
  19. 19.
    Jakoby WB, Ketterer B, Mannervik B: Glutathione transferases: nomenclature. Biochem Pharmacol 33: 2539–2540 (1984).PubMedGoogle Scholar
  20. 20.
    Jepson I, Lay VJ, Holt DC, Bright SWJ, Greenland AJ: Cloning and characterization of maize herbicide safener-induced cDNAs encoding subunits of glutathione S-transferase isoforms I, II and IV. Plant Mol Biol 26: 1855–1866 (1994).PubMedGoogle Scholar
  21. 21.
    Kolm RH, Danielson UH, Zhang Y, Talalay P, Mannervik B: Isothiocyanates as substrates for human glutathione transferases: structure-activity studies. Biochem J 311: 453–459 (1995).PubMedGoogle Scholar
  22. 22.
    Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685 (1970).PubMedGoogle Scholar
  23. 23.
    Marrs KA: The functions and regulation of glutathione Stransferases in plants. Annu Rev Plant Physiol Plant Mol Biol 47: 127–158 (1996).PubMedGoogle Scholar
  24. 24.
    Marrs KA, Alfenito MR, Lloyd AM, Walbot V: A glutathione S-transferase involved in vacuolar transfer encoded by the maize gene Bronze-2. Nature 375: 397–400 (1995).CrossRefPubMedGoogle Scholar
  25. 25.
    O'Connell KM, Breaux EJ, Fraley RT: Different rates of metabolism of two chloroacetanilide herbicides in Pioneer 3320 corn. Plant Physiol 86: 359–363 (1988).Google Scholar
  26. 26.
    Rossini L, Jepson I, Greenland AJ, Sari Gorla M: Characterization of glutathione S-transferase isoforms in three maize inbred lines exhibiting differential sensitivity to alachlor. Plant Physiol 112: 1595–1600 (1996).PubMedGoogle Scholar
  27. 27.
    Rossini L, Pé ME, Frova C, Hein K, Sari-Gorla M: Molecular analysis andmapping of two genes encoding maize glutathione S-transferases (GST I and GST II). Mol Gen Genet 248: 535–539 (1995).PubMedGoogle Scholar
  28. 28.
    Sambrook J, Fritch EF, Maniatis T: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1989).Google Scholar
  29. 29.
    Takahashi Y, Nagata T: parB: An auxin-regulated gene encoding glutathione S-transferase. Proc Natl Acad Sci USA 89: 56–59 (1992).PubMedGoogle Scholar
  30. 30.
    Timmerman KP: Molecular characterization of corn glutathione S-transferase isozymes involved in herbicide detoxication. Physiol Plant 77: 465–471 (1989).Google Scholar
  31. 31.
    Wendel A: Glutathione peroxidase. Meth Enzymol 77: 325–333 (1981).PubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 1998

Authors and Affiliations

  • David P. Dixon
    • 1
  • David J. Cole
    • 2
  • Robert Edwards
    • 1
  1. 1.Crop Protection Group, Department of Biological SciencesUniversity of DurhamDurhamUK
  2. 2.Rhône-Poulenc Agriculture Ltd.EssexUK

Personalised recommendations