Abstract β-mannans are plant structural and storage polysaccharides prevalent in the human diet. Their degradation in the gastrointestinal tract is mediated by the human gut microbiota (HGM) through expression of a plethora of carbohydrate-active enzymes (CAZymes), although our understanding of the details of mannan breakdown is lacking. In this study, a prominent HGM member, Bacteroides cellulosilyticus (Bc), was found to be exceptionally efficient at utilising β-mannans, mediated by the expression of a single polysaccharide utilisation locus (PUL). Amongst the predicted surface CAZymes encoded in the PUL, we identified a family 26 glycoside hydrolase of an unusual molecular architecture. Bc WH2GH26 contains a putative carbohydrate-binding module (CBM) directly intercalated into its catalytic domain, unlike classical CBMs which are located at the N- or C-termini of the catalytic domain. Phylogenetic and functional analyses of this internal CBM, and a homologue from another mannan user Bacteroides uniformis, revealed a narrow specificity for β-mannans and support their classification as a novel CBM family, CBM112. To investigate the functional basis for the unusual enzyme architecture, the effect of the CBM on the catalytic activity of the enzyme was assessed. No significant differences in the kinetic parameters were found between the full-length and CBM deletion constructs against both soluble and insoluble mannans. The potential role of the internal CBM in enzyme function is discussed in the context of the likely localisation of the Bc WH2GH26 in the outer membrane utilisome encoded by the Bc mannan PUL.
Łoś et al. (Thu,) studied this question.