Purification and biochemical characterization of an acidic α-galactosidase from the moringa oleifera seeds

Abstract

A non-glycosylated acidic α-galactosidase was purified from Moringa oleifera (M.oleifera) seeds by a combination of ammonium sulphate precipitation, CM-cellulose (carboxymethyl-cellulose), phenyl Sepharose and Sephadex G-150 column chromatography techniques. In gel filtration the protein eluted as a single peak with a native molecular mass of ~66 kDa. The enzyme migrated as a single band in SDS-PAGE that corresponded to ~64 kDa both under reducing and non-reducing conditions. The pH and temperature optimum of the M. oleifera α-galactosidase was found to be pH 5.0 and 50°C respectively. More than 90% of the enzyme activity is stable between pH 2-6 after 24h of incubation. The enzyme was more than 80% stable at 50°C after 1 h of incubation. The enzyme activity was completely inhibited by Ag+, Hg2+, Cu2+ and SDS at 1 mM concentration. EDTA and reducing agents did not show any effect on enzyme activity. The enzyme was found to be localized in protein bodies in the seed.