Purification and Characterization of 1, 2-α-Mannosidase of Aspergillus oryzae

Abstract

1, 2-α-Mannosidase was purified approximately 1, 400-fold from an enzyme product of Aspergillus oryzae. The enzyme showed a single band in disc gel electrophoresis and the molecular weight was estimated to be about 49, 000 daltons by gel exclusion chromatography. The substrate specificity of the enzyme was examined with mannooligosaccharides, yeast mannan, glycopeptides, and a glycoprotein. The α-(1→2)-linking mannose residues located at the nonreducing-ends of the substrates were selectively removed by the enzyme, whereas p-nitrophenyl α-D-mannopyranoside was completely stable to the enzyme. α-(1→2)-Linking mannose residues in intact bovine pancreatic ribonuclease B were also removed completely with the enzyme. The enzyme showed an optimum pH in the range of pH 4.9 to 5.3 and had a K_m value of 0.57mM with 1, 2-α-mannobiose. The present α-mannosidase was quite stable, and the activity was inhibited by D-mannono-γ-lactone and by heavy metal ions, including zinc ions.