Functional characterization of a gene locus from an uncultured gut Bacteroides conferring xylo-oligosaccharides utilization to Escherichia coli

In prominent gut Bacteroides strains, sophisticated strategies have been evolved to achieve the complete degradation of dietary polysaccharides such as xylan, which is one of the major components of the plant cell wall. Polysaccharide Utilization Loci (PULs) consist of gene clusters encoding different proteins with a vast arsenal of functions, including carbohydrate binding, transport and hydrolysis. Transport is often attributed to TonB-dependent transporters, although major facilitator superfamily (MFS) transporters have also been identified in some PULs. However, until now, few of these transporters have been biochemically characterized. Here, we targeted a PUL-like system from an uncultivated Bacteroides species that is highly prevalent in the human gut metagenome. It encodes three glycoside-hydrolases specific for xylo-oligosaccharides, a SusC/SusD tandem homolog and a MFS transporter. We combined PUL rational engineering, metabolic and transcriptional analysis in Escherichia coli to functionally characterize this genomic locus. We demonstrated that the SusC and the MFS transporters are specific for internalization of linear xylo-oligosaccharides of polymerization degree up to 3 and 4 respectively. These results were strengthened by the study of growth dynamics and transcriptional analyses in response to XOS induction of the PUL in the native strain, Bacteroides vulgatus