Understanding the adhesion mechanism of a mucin binding domain from Lactobacillus fermentum and its role in enteropathogen exclusion

Lactobacillus species possesses surface exposed Mucin Binding Protein (MucBP) which plays a role in adhesion to gastrointestinal mucin. MucBP contains one or more mucin binding domain (MBD), the functionality of which has yet not been characterized thoroughly. Here, we have characterized a 93-amino acid MBD (MBD_{93}) of MucBP (LAF_0673) from Lactobacillus fermentum. Multiple sequence alignment of L. fermentum MBD_{93} exhibited ∼60% sequence homology with MBDs from other Lactobacillus species. Further, we cloned, expressed and purified MBD_{93} from Escherichia coli as N-terminal histidine-tagged protein (6X His-MBD_{93}). The purified MBD_{93} was able to bind to mucin and showed strong affinity towards the terminally expressed mucin glycans viz. N-acetylgalactosamine (GalNAc), N-acetylglucosamine (GlcNAc), Galactose (Gal), and Sialic acid (N-acetylneuraminic acid; Neu5Ac). In silico experiments further confirmed the interaction between homology modeled MBD_{93} to mucin glycans through hydrogen-bonding with its surface amino acid residues Ser^{57}, Pro^{58}, Ile^{60}, Tyr^{63} and Ala^{65}. We also have demonstrated that MBD_{93} was able to inhibit the adhesion of enteric pathogens, including E. coli, Salmonella Paratyphi A, Shigella sonnei and Proteus vulgaris to mucin. Our results suggested that L. fermentum MBD_{93} is a functionally sufficient unit to act as an adhesin and to protect from invading enteric pathogens.