Purification and characterization of 1,2-alpha-mannosidase of Aspergillus oryzae

1,2-alpha-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 alpha-(1 leads to 2)-linking mannose residues located at the nonreducing-ends of the substrates were selectively removed by the enzyme, whereas p-nitrophenyl alpha-D-mannopyranoside was completely stable to the enzyme. alpha-(1 leads to 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 Km value of 0.57 mM with 1,2-alpha-mannobiose. The present alpha-mannosidase was quite stable, and the activity was inhibited by D-mannono-gamma-lactone and by heavy metal ions, including zinc ions.