Threonine synthase of Lemna paucicostata hegelm. 6746

Abstract

Threonine synthase (TS) was purfied approximately 40-fold from Lemna pacicostata, and some of its properties determined by use of a sensitive and spedic assay. During the core of its purification, TS was separated from cystathionine γ-synthase, establishing the separte identity of these enzymes. Compared to cystathionine γ-synthase, TS is relatively insensitive to irreversible inhibition by propargylgycine (both in vitro and in vivo) and to gabaculine, vinylglycine, or cysteine in vitro. TS is highly specific for O-phospho-L-homoserine (OPH) and water (hydroxyl ion). Nucleophilic attack by hydroxyl ion is restricted to carbon-3 ofOPH and proceeds sterospecifically to form threonine rather than allo-threonine. The K_m for OPH, determined at saturating Sadenosylmethioine (AdoMet), is 2.2 to 6.9 micromolar, two orders of maugtude less than values reported for TS from other plant tissues. AdoMet markedly stimultes the enzyme in a reversible and cooperative manner, consistent with its proposed role in regulation of methioine biosynthesis. Cysteine (1 milar) caused a sLight (26%) reversible inhibition of the enzyme. Activities of TS isolated from Lemna were inversely related to the methionine nutrition of the plants. Down-regulation ofTS by methioni may help to limit the overproduction ofthreone that could result from allosteric stimulation of the enzyme by AdoMet. No evidence was obtaied for feedback inhibition, or covalent modification of TS by threonine and/or isolencine.