Purification and characterization of beta-N-acetylgalactosaminidase from Bacillus sp. AT173-1

Beta-N-Acetylgalactosaminidase [EC 3.2.1.53] was purified to homogeneity from the culture media of Bacillus sp. AT173-1. The enzyme has a molecular weight of 48,000 as estimated by SDS-PAGE under reducing conditions and an isoelectric point of 4.3. The enzyme requires dithiothreitol as an activator and is most active at pH 6.0. Analysis of its substrate specificity using 2-aminopyridine-labeled oligosaccharides as substrates revealed the enzyme specifically hydrolyzes beta-N-acetylgalactosaminyl linkages of GalNAcbeta1-4Galbeta1-4Glc, GalNAcbeta1-3Gal alpha1-4Galbeta1-4Glc, and N-glycans terminating with beta-N-acetylgalactosamine residues but not those with beta-N-acetylglucosamine residues. The enzyme is thus a novel beta-N-acetylgalactosaminidase with practically no beta-N-acetylglucosaminidase activity.