Abstract: 1-aminocyclopropane-1-carboxylate (ACC) synthase is a principal enzyme that catalyses the committed step in phytohormone ethylene biosynthesis. Previous evidence indicates that the hypervariable C-terminus of ACC synthase is most likely to be processed proteolytically in vivo. However, the protease responsible has not been identified thus far. In the present study, we detected proteolytic activity against ACC synthase (LeACS2) in tomato (Lycopersicon esculentum Mill.) fruit extract based on a newly established in vitro assay system. Purification of the protease through DEAE, gel filtration and MonoQ chromatography resulted in considerable enrichment of a 64-kDa protein species. Subsequent biochemical analysis of the purified tomato protease revealed that the optimal conditions for its proteolytic activity were at pH 8.0 and at 37 °C. In addition, the protease activity was blocked completely by the metalloprotease inhibitor 1,10-phenanthroline. The present study represents the first report on the isolation of an ACC synthase-processing protease from plant tissues.

Key words: 1-aminocyclopropane-1-carboxylate (ACC) synthase, biochemical analysis, C-terminal proteolysis, isolation, protease.