Degradation of salivary MUC5B mucin by oral biofilm bacteria

Introduction: Complex substrates, like salivary mucins (MUC5B), require an arsenal of glycosidases and proteases to sequentially degrade the oligosaccharides and polypeptide backbone. In lieu of complex salivary MUC5B, centrifuged and processed saliva or commercially reduced mucin subunits, markedly less complex mixtures than encountered by the oral microflora in vivo, have served as experimental substrates for growth of oral bacteria and utilisation of the substrates as nutrients. MUC5B is a complex oligomeric glycoprotein, heterogeneous in size (1.5-40 mDa), with a repertoire of diverse oligosaccharides in composition and charge. Subpopulations of salivary MUC5B have been identified, differing in both molecular organization and solubility.

Objective: The aim of this study was to determine whether dental biofilm bacteria cooperated in degrading human salivary MUC5B and identify proteolytic degradation of the mucin polypeptide backbone.

Methods: Freshly collected supragingival plaque from human volunteers was incubated with purified human MUC5B. Overall degradation of MUC5B was determined by SDS-PAGE, while glycan degradation was estimated by a lectin-binding assay. The microbial sources of degradative glycosidase and proteolytic activities were investigated by incubating the dental plaque preparations with fluorescent-labeled substrates.

Results: Supragingival plaque bacteria expressed the requisite glycosidases and proteolytic activity needed to degrade complex salivary glycoproteins such as MUC5B. These plaque enzymes degraded the oligosaccharides and the polypeptide backbone of salivary MUC5B. After 8h, lectin binding declined to all external MUC5B constituent monosaccharides; degradation of glcNAc and galNAc was minimal at 8 h consistent with their internal positions in the glycans. Bacterial proteases degraded the central domain of the MUC5B peptide backbone as shown by the appearance of cleavage fragments on SDS-PAGE.

Conclusion: Certain bacteria in dental plaque appear to selectively express glycosidases and proteases, which sequentially degrade complex human salivary glycoproteins.

Supported by The Swedish Research Council (K2005-06X-12266)

and NIH R01DE08590 (MCH).