Partial purification and property of Mg-dechelatase from harvested banana fruit
Banana fruit fail to develop a fully yellow peel when ripening at tropical temperature (>24°C), resulting in “green-ripe” fruit. Our previous study showed that the inhibition of Mg-dechelatase activity in banana peel at 30°C might contribute to a repressed chlorophyll degradation, leading to uneven degreening appearance. In the present study, Mg-dechelatase was partially purified from banana peel tissues by ammonium sulfate fractionation and the property of the enzyme was characterized. A 2.21-fold purification of Mg-dechelatase was obtained using 45-90% ammonium sulfate fractionation. However, Mg-dechelatase lost most of its activity when further subjected to a phenyl-sepharose CL-6B chromatography. High stability of the enzyme was observed at a temperature range of 30-70°C and about 48% of the activity was maintained after incubated at 100°C for 40 min. The optimum reaction temperature of Mg-dechelatase was determined to be 40-50°C. Mg-dechelatase activity gradually increased with increasing pH values at a range of 6.5-9.5. The Km for banana Mg-dechelatase with chlorophyllin as a substrate was 8.09 nmol/L. The activity of the enzyme was strongly inhibited by β-mercaptoethanol and L-ascorbic acid (ASA), but to an extent by Na+, Mg2+, citric acid, reduced glutathione (GSH). However, Fe2+, Ca2+ and H2O2 had a marked activation effect on the banana Mg-dechelatase activity.