The human gut symbiont Ruminococcus gnavus shows specificity to blood group A antigen during mucin glycan foraging: Implication for niche colonisation in the gastrointestinal tract

DOI: 10.1371/journal.pbio.3001498

Authors: Haiyang Wu (Quadram Institute Bioscience) , Emmanuelle H. Crost (Quadram Institute Bioscience) , C. David Owen (Diamond Light Source; Research Complex at Harwell) , Wouter Van Bakel (Quadram Institute Bioscience) , Ana Martínez Gascueña (Quadram Institute Bioscience) , Dimitrios Latousakis (Quadram Institute Bioscience) , Thomas Hicks (Quadram Institute Bioscience) , Samuel Walpole (University of East Anglia) , Paulina A. Urbanowicz (Ludger Ltd) , Didier Ndeh (Quadram Institute Bioscience) , Serena Monaco (University of East Angli) , Laura Sánchez Salom (Quadram Institute Bioscience) , Ryan Griffiths (Quadram Institute Bioscience) , Raven S. Reynolds (Quadram Institute Bioscience) , Anna Colvile (Diamond Light Source) , Daniel I. R. Spencer (Ludger Ltd) , Martin Walsh (Diamond Light Source) , Jesus Angulo (University of East Anglia) , Nathalie Juge (Quadram Institute Bioscience)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Plos Biology , VOL 19

State: Published (Approved)
Published: December 2021

Abstract: The human gut symbiont Ruminococcus gnavus displays strain-specific repertoires of glycoside hydrolases (GHs) contributing to its spatial location in the gut. Sequence similarity network analysis identified strain-specific differences in blood-group endo-β-1,4-galactosidase belonging to the GH98 family. We determined the substrate and linkage specificities of GH98 from R. gnavus ATCC 29149, RgGH98, against a range of defined oligosaccharides and glycoconjugates including mucin. We showed by HPAEC-PAD and LC-FD-MS/MS that RgGH98 is specific for blood group A tetrasaccharide type II (BgA II). Isothermal titration calorimetry (ITC) and saturation transfer difference (STD) NMR confirmed RgGH98 affinity for blood group A over blood group B and H antigens. The molecular basis of RgGH98 strict specificity was further investigated using a combination of glycan microarrays, site-directed mutagenesis, and X-ray crystallography. The crystal structures of RgGH98 in complex with BgA trisaccharide (BgAtri) and of RgGH98 E411A with BgA II revealed a dedicated hydrogen network of residues, which were shown by site-directed mutagenesis to be critical to the recognition of the BgA epitope. We demonstrated experimentally that RgGH98 is part of an operon of 10 genes that is overexpresssed in vitro when R. gnavus ATCC 29149 is grown on mucin as sole carbon source as shown by RNAseq analysis and RT-qPCR confirmed RgGH98 expression on BgA II growth. Using MALDI-ToF MS, we showed that RgGH98 releases BgAtri from mucin and that pretreatment of mucin with RgGH98 confered R. gnavus E1 the ability to grow, by enabling the E1 strain to metabolise BgAtri and access the underlying mucin glycan chain. These data further support that the GH repertoire of R. gnavus strains enable them to colonise different nutritional niches in the human gut and has potential applications in diagnostic and therapeutics against infection.

Journal Keywords: Blood groups; Mucin; NMR spectroscopy; Polymerase chain reaction; Operons; Crystal structure; Nuclear magnetic resonance; Gut bacteria

Diamond Keywords: Bacteria; Gut Microbiota; Enzymes

Subject Areas: Biology and Bio-materials, Medicine


Instruments: I03-Macromolecular Crystallography , I04-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography , VMXi-Versatile Macromolecular Crystallography in situ

Added On: 28/12/2021 16:35

Documents:
journal.pbio.3001498.pdf

Discipline Tags:

Health & Wellbeing Structural biology Drug Discovery Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)