Three-dimensional structure of glutathione S-transferase from Arabidopsis thaliana at 2.2 A resolution: structural characterization of herbicide-conjugating plant glutathione S-transferases and a novel active site architecture

J Mol Biol. 1996 Jan 19;255(2):289-309. doi: 10.1006/jmbi.1996.0024.

Abstract

Glutathione S-transferases (GST) are a family of multifunctional enzymes involved in the metabolization of a broad variety of xenobiotics and reactive endogenous compounds. The interest in plant glutathione S-transferases may be attributed to their agronomic value, since it has been demonstrated that glutathione conjugation for a variety of herbicides is the major resistance and selectivity factor in plants. The three-dimensional structure of glutathione S-transferase from the plant Arabidopsis thaliana has been solved by multiple isomorphous replacement and multiwavelength anomalous dispersion techniques at 3 A resolution and refined to a final crystallographic R-factor of 17.5% using data from 8 to 2.2 A resolution. The enzyme forms a dimer of two identical subunits each consisting of 211 residues. Each subunit is characterized by the GST-typical modular structure with two spatially distinct domains. Domain I consists of a central four-stranded beta-sheet flanked on one side by two alpha-helices and on the other side by an irregular segment containing three short 3(10)-helices, while domain II is entirely helical. The dimeric molecule is globular with a prominent large cavity formed between the two subunits. The active site is located in a cleft situated between domains I and II and each subunit binds two molecules of a competitive inhibitor S-hexylglutathione. Both hexyl moieties are oriented parallel and fill the H-subsite of the enzyme's active site. The glutathione peptide of one inhibitor, termed productive binding, occupies the G-subsite with multiple interactions similar to those observed for other glutathione S-transferases, while the glutathione backbone of the second inhibitor, termed unproductive binding, exhibits only weak interactions mediated by two polar contacts. A most striking difference from the mammalian glutathione S-transferases, which share a conserved catalytic tyrosine residue, is the lack of this tyrosine in the active site of the plant glutathione S-transferase.

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / enzymology*
  • Binding Sites
  • Crystallography, X-Ray
  • Enzyme Inhibitors / metabolism
  • Enzyme Inhibitors / pharmacology
  • Glutathione / analogs & derivatives
  • Glutathione / metabolism
  • Glutathione / pharmacology
  • Glutathione Transferase / antagonists & inhibitors
  • Glutathione Transferase / chemistry*
  • Glutathione Transferase / metabolism
  • Herbicides / metabolism
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Conformation*
  • Protein Structure, Secondary
  • Sequence Alignment

Substances

  • Enzyme Inhibitors
  • Herbicides
  • Glutathione Transferase
  • Glutathione
  • hexylglutathione