Extraction and Properties of 1-Aminocyclopropane-1-Carboxylate Synthase in Banana Fruit.

Abstract

The determination of 1-aminocyclopropane-1-carboxylate (ACC) synthase activity is essential in understanding its role in the ethylene biosynthesis during fruit ripening. Because of the high level of soluble tannins, to date, there has been no report on successful determination of banana ACC synthase activity. In this study, we examined the method of Badran and Jones (1965) for the extraction of ACC synthase from banana fruit. The extraction procedure consists of homogenizing the pulp in polyethyleneglycol (PEG)-added buffer, and then washing the homogenate with acetone. This PEG-acetone method gave a high ACC synthase activity. The common extraction method using polyvinylpyrrolidone (PVP) yielded a much lower or no ACC synthase activity. The changes in ACC synthase activity determined by the PEG-acetone method correlated with changes in ethylene production during ripening. The optimum pH, Km value for substrate S-adenosylmethionine (SAM), and half-life in the presence of SAM for banana ACC synthase were 9.0, 88μM, and 18 min respectively. These values were within the range previously reported for ACC synthase in various plant tissues. From these results, we recommend the following extraction method to determine banana ACC synthase activity : freeze the flesh tissue in liquid nitrogen, store it at -80°C until use, homogenize the frozen sample in a Waring blender with PEG-added extraction buffer, and later precipitate the enzyme mixture with acetone.