A 2,4-D-INDUCIBLE GLUTATHIONE-S-TRANSFERASE FROM SOYBEAN (GLYCINE-MAX) - PURIFICATION, CHARACTERIZATION AND INDUCTION

Glutathione S-transferases (GSTs; EC 2.5.1.18) have recently been proposed to form one large group among the auxin-induced proteins. However, the properties and regulation of such auxin-responsive GSTs in the plant still await detailed investigation. In this study, a 2,4-dichloro-phenoxyacetic acid (2,4-D)-inducible GST isozyme from soybean (Glycine mar [L.] Merr. cv. Williams) was purified to near homogeneity by anion-exchange and affinity chromatography on S-hexylglutathione agarose. The native enzyme had a molecular mass of 49 kDa, as determined by gel filtration, and consisted of 26-kDa subunits. The purified GST conjugated glutathione to 1-chloro-2,4-dinitrobenzene and to the herbicide metolachlor, but not to the other GST substrates atrazine, fluorodifen or trans-cinnamic acid. The N-terminal amino acid sequence shared significant homology with the deduced polypeptide sequences of two 2,4-D-inducible genes from tobacco, parA and CNT107. The levels of the 26-kDa GST subunit protein in soybean hypocotyls were analysed by immunoblotting. At micromolar concentrations, 2,4-D induced a transient increase in net accumulation of GST, whereas indole-3-acetic acid or 1-naphthaleneacetic acid did not increase the GST levels. Known inhibitors of polar auxin transport, including 2,3 ,5-tri-iodobenzoic acid, N-1-naphthylphtharamic acid and analogues thereof, differed widely in their ability to elicit GST protein accumulation. It is concluded that the induction of soybean GST by 2,4-D and by some of the auxin transport inhibitors is not related to auxin activity or to changes in the endogenous auxin levels.