Dataset Information

Biochemical Basis of Xylooligosaccharide Utilisation by Gut Bacteria.

ABSTRACT: Xylan is one of the major structural components of the plant cell wall. Xylan present in the human diet reaches the large intestine undigested and becomes a substrate to species of the gut microbiota. Here, we characterised the capacity of Limosilactobacillus reuteri and Blautia producta strains to utilise xylan derivatives. We showed that L. reuteri ATCC 53608 and B. producta ATCC 27340 produced β-D-xylosidases, enabling growth on xylooligosaccharide (XOS). The recombinant enzymes were highly active on artificial (p-nitrophenyl β-D-xylopyranoside) and natural (xylobiose, xylotriose, and xylotetraose) substrates, and showed transxylosylation activity and tolerance to xylose inhibition. The enzymes belong to glycoside hydrolase family 120 with Asp as nucleophile and Glu as proton donor, as shown by homology modelling and confirmed by site-directed mutagenesis. In silico analysis revealed that these enzymes were part of a gene cluster in L. reuteri but not in Blautia strains, and quantitative proteomics identified other enzymes and transporters involved in B. producta XOS utilisation. Based on these findings, we proposed a model for an XOS metabolism pathway in L. reuteri and B. producta strains. Together with phylogenetic analyses, the data also revealed the extended xylanolytic potential of the gut microbiota.

SUBMITTER: Singh RP

PROVIDER: S-EPMC8954004 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Publications

Biochemical Basis of Xylooligosaccharide Utilisation by Gut Bacteria.

Singh Ravindra Pal RP Bhaiyya Raja R Thakur Raksha R Niharika Jayashree J Singh Chandrajeet C Latousakis Dimitrios D Saalbach Gerhard G Nepogodiev Sergey A SA Singh Praveen P Sharma Sukesh Chander SC Sengupta Shantanu S Juge Nathalie N Field Robert A RA

International journal of molecular sciences 20220310 6

Xylan is one of the major structural components of the plant cell wall. Xylan present in the human diet reaches the large intestine undigested and becomes a substrate to species of the gut microbiota. Here, we characterised the capacity of Limosilactobacillus reuteri and Blautia producta strains to utilise xylan derivatives. We showed that L. reuteri ATCC 53608 and B. producta ATCC 27340 produced β-D-xylosidases, enabling growth on xylooligosaccharide (XOS). The recom ...[more]

PMID: 35328413

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Project description:In the human gut microbiota, Bacteroidetes break down dietary and endogenous glycosides through highly specific polysaccharide utilization loci (PULs). PULs encode a variety of sensor regulators, binding proteins, transporters, and carbohydrate-active enzymes (CAZymes). Surface glycan-binding proteins (SGBPs) are essential for the efficient capture of the glycosides present on the cell surface, providing Bacteroidetes with a competitive advantage in colonizing their habitats. Here, we present the functional and structural characterization of a SusD-like protein encoded by a xylooligosaccharide (XOS) PUL from an uncultured human gut Bacteroides strain. This locus is also conserved in Bacteroides vulgatus, thereby providing new mechanistic insights into the role of SGBPs in the metabolism of dietary fiber of importance for gut health. Various in vitro analyses, including saturation transfer difference nuclear magnetic resonance (STD-NMR) spectroscopy, revealed that the SusD-like protein cannot bind to the cognate substrate of the XOS PUL, although its presence is essential for the PUL to function. Analysis of the crystal structure of the SusD-like protein reveals an unfolded binding surface and the absence or inappropriate orientation of several key residues compared with other known SusD-like structures. These results highlight the critical role of the SusD-like protein in the transport of oligosaccharides and provide fundamental knowledge about the structure-function of SusC/D-like transporters, revealing that the binding specificity of SusD-like SGBPs does not necessarily reflect the uptake specificity of the transporter. IMPORTANCE The metabolization of dietary fiber is a crucial function for many gut bacteria, especially Bacteroidetes, which are particularly well adapted for recognizing, binding, transporting, and degrading glycosides. In this study, we report the functional and structural characterization of a SusD-like protein involved in xylooligosaccharide utilization by an uncultured gut Bacteroides strain. We demonstrate that while this protein is structurally similar to many canonical Bacteroidetes surface glycan-binding proteins, it cannot bind the substrate taken up by the cognate SusC-like transporter. This lack of binding might be explained by the absence of several key residues known to be involved in oligosaccharide binding and/or the possible necessity of the SusC-like protein to be present to create a cooperative binding site. The term "surface glycan-binding proteins" generally used for SusD-like proteins is thus not generic. Overall, this study allowed us to revisit the concept of glycoside utilization by Bacteroidetes, in particular those strains that feed on the short fibers naturally present in some dietary compounds or on the leftovers of other microbes.

Dataset Information

Biochemical Basis of Xylooligosaccharide Utilisation by Gut Bacteria.

Publications

Biochemical Basis of Xylooligosaccharide Utilisation by Gut Bacteria.

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