Molecular and catalytic characterization of a phi class glutathione transferase from Cathaya argyrophylla

Plant phi class glutathione transferases (GSTs) play important roles in stress tolerance and detoxification metabolism. This study reports the cloning, expression and biochemical characteristics of a phi GST gene (CaGSTF) from the endemic and endangered conifer Cathaya argyrophylla. The recombinant CaGSTF showed GSH-conjugating activity towards the substrate NBD-Cl and CDNB. Kinetic analysis revealed low catalytic efficiency with a kcat/KmGSH value of 9.82 mM−1S−1. The CaGSTF proved to be a thermolabile enzyme, at 40 °C the enzyme’s activity was nearly abolished. Site-directed mutagenesis revealed that Ser12, Lys42, Ile55, Glu67 and Ser68 of CaGSTF are critical components of glutathione-binding sites that contribute to the enzyme’s catalytic activity. Compared to other plant phi GSTs and conifer tau GSTs, CaGSTF showed a narrow substrate spectrum, low catalytic efficiency and thermolability. These atypical properties suggest the enzyme may have a limited functional role in the organism’s adaptation to environmental stresses in the subtropical regions.

► We cloned a new phi class GST (CaGSTF) from the endangered conifer Cathaya argyrophylla. ► The CaGSTF showed a narrow substrate spectrum, low catalytic efficiency and thermolability. ► Ser12, Lys42, Ile55, Glu67 and Ser68 of CaGSTF were identified as glutathione-binding residues. ► Glutathione-binding residues of CaGSTF are critical for the enzyme’s catalytic activity.