α-N-Acetylgalactosaminidase from Infant-associated Bifidobacteria Belonging to Novel Glycoside Hydrolase Family 129 Is Implicated in Alternative Mucin Degradation Pathway

Bifidobacteria inhabit the lower intestine of mammals including humans where the mucin gel layer forms a space for commensal bacteria. We previously identified that infant-associated bifidobacteria possess an extracellular membrane-bound endo-alpha-N-acetylgalactosaminidase (EngBF) that may be involved in degradation and assimilation of mucin-type oligosaccharides. However, EngBF is highly specific for core-1-type O-glycan (Gal beta 1-3GalNAc alpha 1-Ser/Thr), also called T antigen, which is mainly attached onto gastroduodenal mucins. By contrast, core-3-type O-glycans (GlcNAc beta 1-3GalNAc alpha 1-Ser/Thr) are predominantly found on the mucins in the intestines. Here, we identified a novel alpha-N-acetylgalactosaminidase (NagBb) from Bifidobacterium bifidum JCM 1254 that hydrolyzes the Tn antigen (GalNAc alpha 1-Ser/Thr). Sialyl and galactosyl core-3 (Gal beta 1-3/4GlcNAc beta 1-3(Neu5Ac alpha 2-6)GalNAc alpha 1-Ser/Thr), a major tetrasaccharide structure on MUC2 mucin primarily secreted from goblet cells in human sigmoid colon, can be serially hydrolyzed into Tn antigen by previously identified bifidobacterial extracellular glycosidases such as alpha-sialidase (SiaBb2), lacto-N-biosidase (LnbB), beta-galactosidase (BbgIII), and beta-N-acetylhexosaminidases (BbhI and BbhII). Because NagBb is an intracellular enzyme without an N-terminal secretion signal sequence, it is likely involved in intracellular degradation and assimilation of Tn antigen-containing polypeptides, which might be incorporated through unknown transporters. Thus, bifidobacteria possess two distinct pathways for assimilation of O-glycans on gastroduodenal and intestinal mucins. NagBb homologs are conserved in infant-associated bifidobacteria, suggesting a significant role for their adaptation within the infant gut, and they were found to form a new glycoside hydrolase family 129.