Characterization of ornithine decarboxylase from Pseudomonas syringae pv. phaseolicola and its inhibition by phaseolotoxin

Abstract

Pseudomonas syringae pv. phaseolicola, the causative agent of halo blight of French bean ( Phaseolus vulgaris L.), produces phaseolotoxin (PT), which functions as a chlorosis-inducing extracellular phytotoxin by inhibiting ornithine carbamoyltransferase (OCT). We previously demonstrated that PT also inhibits ornithine decarboxylase (ODC), a key enzyme in polyamine biosynthesis, in preparations of bean leaves. In this study, the ODC enzyme was isolated from PT-producing bacterial cultures to study enzyme characteristics and to assess the effect of PT on ODC activity. The enzyme was purified using a four-step procedure comprised of ammonium sulfate precipitation, gel filtration chromatography, ion-exchange chromatography and adsorption to hydroxylapatite. ODC was optimally active at pH 7.3 and showed hyperbolic kinetics with a K m of 0.19±0.04 mM for ornithine. PT inhibited the ODC enzyme with a K i of 16±0.05 μM. In addition, the specific suicide inhibitor DFMO also inhibited ODC with a K i of 26±0.2 μM.