Characterization of the Streptococcus pneumoniae BgaC Protein as a Novel Surface β-Galactosidase with Specific Hydrolysis Activity for the Galβ1-3GlcNAc Moiety of Oligosaccharides

Streptococcus pneumoniae is a causative agent of high morbidity and mortality. Although sugar moieties have been recognized as ligands for initial contact with the host, only a few exoglycosidases have been reported to occur inS. pneumoniae . In this study, a putative β-galactosidase, encoded by thebgaC gene ofS. pneumoniae , was characterized for its enzymatic activity and virulence. The recombinant BgaC protein, expressed and purified fromEscherichia coli , was found to have a highly regiospecific and sugar-specific hydrolysis activity for the Galβ1-3-GlcNAc moiety of oligosaccharides. Interestingly, the BgaC hydrolysis activity was localized at the cell surface ofS. pneumoniae , indicating that BgaC is expressed as a surface protein although it does not have a typical signal sequence or membrane anchorage motif. The surface localization of BgaC was further supported by immunofluorescence microscopy analysis using an antibody raised against BgaC and by a reassociation assay with fluorescein isothiocyanate-labeled BgaC. Although thebgaC deletion mutation did not significantly attenuate the virulence ofS. pneumoniae in vivo, thebgaC mutant strain showed relatively low numbers of viable cells compared to the wild type after 24 h of infection in vivo, whereas the mutant showed higher colonization levels at 6 and 24 h postinfection in vivo. Our data strongly indicate for the first time thatS. pneumoniae bgaC encodes a surface β-galactosidase with high substrate specificity that is significantly associated with the infection activity of pneumococci.