Comparative Analysis of Two Stress-Inducible tau Class Glutathione Transferases from Glycine max Revealed Significant Catalytic and Structural Diversification

Protein Pept Lett. 2017;24(10):922-935. doi: 10.2174/0929866524666171026125300.

Abstract

Background: Glutathione transferases (GSTs, EC. 2.5.1.18) form a large group of multifunctional enzymes that are involved in the metabolism and inactivation of a wide range of endogenous and xenobiotic compound as well as in cell regulation and response to several biotic and abiotic stresses.

Objectives: In the present work, we report the comparative analysis of the structural and functional features of two isoenzymes (GmGSTU5-5 and GmGSTU8-8) of the glutathione transferase (GST) family from Glycine max.

Methods: Full-length cDNA clones of GmGSTU5-5 and GmGSTU8-8 were derived from RT-PCR of RNA isolated from soybean seedlings and were cloned into a T7 expression vector. Τhe recombinant enzymes were expressed in E. coli and purified by affinity chromatography. Substrate specificity, kinetic and inhibition analysis were carried out towards a range of different xenobiotic compounds and GSH analogues. The thermal stability of the enzymes was also evaluated using activity assays and differential scanning fluorimetry.

Results: Analysis of substrate specificity using a range of thiol substrates and electrophilic compounds suggested that both isoenzymes display broad and overlapping specificities. They are capable of detoxifying major stress-induced toxic products. Study of their ligandin-binding properties by kinetic analysis and molecular modelling indicated that both GmGSTU5-5 and GmGSTU8-8 bind a range of secondary metabolites and plant hormones, suggesting a role in transport or storage of bioactive compounds. Thermostability analysis showed that GmGSTU5-5 and GmGSTU8-8 display extraordinary thermal stability, compared to other plant GSTs.

Conclusion: Our results suggest that GmGSTU5-5 and GmGSTU8-8 display different or overlapping substrate specificities and kinetic properties. The biological role of GmGSTU5-5 and GmGSTU8-8 may be relevant to the detoxification of toxic compounds or the binding of bioactive metabolites that function in cell regulation and stress defence mechanisms.

Keywords: Abiotic stress; Glycine max; biotic stress; glutathione transferase; isoenzymes; xenobiotic metabolism.

Publication types

  • Comparative Study

MeSH terms

  • Amino Acid Sequence
  • Catalysis
  • Escherichia coli
  • Glutathione Transferase / chemistry
  • Glutathione Transferase / metabolism*
  • Glycine max / chemistry*
  • Kinetics
  • Models, Molecular
  • Plant Growth Regulators / metabolism
  • Plant Proteins / chemistry
  • Plant Proteins / metabolism*
  • Protein Binding
  • Protein Structure, Secondary
  • Seedlings
  • Stress, Physiological
  • Substrate Specificity
  • tau Proteins / chemistry
  • tau Proteins / metabolism*

Substances

  • Plant Growth Regulators
  • Plant Proteins
  • tau Proteins
  • Glutathione Transferase