Purification and biochemical characterization of a new strictly ?1,2-mannosidase from breadfruit (Artocarpus communis) seeds.

mannosidases importance in N-glycoproteins processing and degradation prompted us to search for a new source. Thus, one ?-mannosidase was purified from matured breadfruit (Artocarpus communis) seeds, by successive chromatography on DEAE-Sepharose CL-6B, CM-Sepharose CL-6B and Phenyl-Sepharose HP to apparent homogeneity. The enzyme had native molecular weight of approximately 46 kDa. SDS-PAGE analysis resolved a single protein band with molecular weight estimated to 46 kDa. Breadfruit ?-mannosidase had optimal pH (4.6) and temperature (55 °C). Its activity was enhanced in presence of detergents such as Hexansulfonic acid sodium salt, Polyoxyethylen-9-lauryl ether, Nonidet P40, Triton X-100 as well as Ca2+ and Zn2+. The effect of ?-mannosidase inhibitors on the enzyme showed that swainsonine (SW) and 1,4-dideoxy-1,4-imino mannitol (DIM) inhibited it’s hydrolytic activity at low concentrations, while kifunensine (KIF) and deoxymannojirimycin (DMNJ) had no significant effect at the same concentrations on this enzyme. Substrate specificity tests revealed that the enzyme exhibited only ?1,2-mannosidase activity. Thus, breadfruit ?-mannosidase may be a useful tool for applications requiring selective removal of mannose. Since breadfruit seed ?-mannosidase was not sensitive to KIF and cleaved only ?-(1,2) linked mannobiose, this enzyme would belong to class I ?-mannosidases and seems to be ?1,2- mannosidase Kifunensine resistant-like.