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Multivalency plays a major role in biological processes and particularly in the relationship between pathogenic microorganisms and their host that involves protein–glycan recognition. These interactions occur during the first steps of infection, for specific recognition between host and bacteria, but also at different stages of the immune response. The search for high-affinity ligands for studying such interactions involves the combination of carbohydrate head groups with different scaffolds and linkers generating multivalent glycocompounds with controlled spatial and topology parameters. By interfering with pathogen adhesion, such glycocompounds including glycopolymers, glycoclusters, glycodendrimers and glyconanoparticles have the potential to improve or replace antibiotic treatments that are now subverted by resistance. Multivalent glycoconjugates have also been used for stimulating the innate and adaptive immune systems, for example with carbohydrate-based vaccines. Bacteria present on their surfaces natural multivalent glycoconjugates such as lipopolysaccharides and S-layers that can also be exploited or targeted in anti-infectious strategies.

Plant pathogens, like animal ones, use protein-carbohydrate interactions in their strategy for host recognition, attachment, and invasion. The bacterium Ralstonia solanacearum, which is distributed worldwide and causes lethal wilt in many agricultural crops, was shown to produce a potent L-fucose-binding lectin, R. solanacearum lectin, a small protein of 90 amino acids with a tandem repeat in its amino acid sequence. In the present study, surface plasmon resonance experiments conducted on a series of oligosaccharides show a preference for binding to alpha Fuc1-2Gal and alpha Fuc1-6Gal epitopes. Titration microcalorimetry demonstrates the presence of two binding sites per monomer and an unusually high affinity of the lectin for alpha Fuc1-2Gal-containing oligosaccharides (K-D = 2.5 x 10(-7) M for 2-fucosyllactose). R. solanacearum lectin has been crystallized with a methyl derivative of fucose and with the highest affinity ligand, 2-fucosyllactose. X-ray crystal structures, the one with alpha-methyl-fucoside being at ultrahigh resolution, reveal that each monomer consists of two small four-stranded anti-parallel beta-sheets. Trimerization through a 3-fold or pseudo-3-fold axis generates a six-bladed beta-propeller architecture, very similar to that previously described for the fungal lectin of Aleuria aurantia. This is the first report of a beta-propeller formed by oligomerization and not by sequential domains. Each monomer presents two fucose binding sites, resulting in six symmetrically arranged sugar binding sites for the beta-propeller. Crystals were also obtained for a mutated lectin complexed with a fragment of xyloglucan, a fucosylated polysaccharide from the primary cell wall of plants, which may be the biological target of the lectin., QC 20230602

Since the introduction of structural genomics, the protein has been recognized as the most important variable in crystallization. Recent strategies to modify a protein to improve crystal quality have included rationally engineered point mutations, truncations, deletions and fusions. Five naturally occurring variants, differing in 1 - 18 amino acids, of the 177-residue lectin domain of the F17G fimbrial adhesin were expressed and purified in identical ways. For four out of the five variants crystals were obtained, mostly in non-isomorphous space groups, with diffraction limits ranging between 2.4 and 1.1 angstrom resolution. A comparative analysis of the crystal-packing contacts revealed that the variable amino acids are often involved in lattice contacts and a single amino-acid substitution can suffice to radically change crystal packing. A statistical approach proved reliable to estimate the compatibilities of the variant sequences with the observed crystal forms. In conclusion, natural variation, universally present within prokaryotic species, is a valuable genetic resource that can be favourably employed to enhance the crystallization success rate with considerably less effort than other strategies., QC 20230602

SmI2/H2O/amine provides selective cleavage of unsubstituted allyl ethers in good to excellent yields. This method is therefore useful in deprotection of alcohols and carbohydrates., QC 20230602

The F17-G adhesin at the tip of flexible F17 fimbriae of enterotoxigenic Escherichia coli mediates binding to N-acetyl-beta-D-glucosamine-presenting receptors on the microvilli of the intestinal epithelium of ruminants. We report the 1.7 Angstrom resolution crystal structure of the lectin domain of F17-G, both free and in complex with N-acetylglucosamine. The monosaccharide is bound on the side of the ellipsoid-shaped protein in a conserved site around which all natural variations of F17-G are clustered. A model is proposed for the interaction between F17-fimbriated E. coli and microvilli with enhanced affinity compared with the binding constant we determined for F17-G binding to N-acetylglucosamine (0.85 mM(-1)). Unexpectedly, the F17-G structure reveals that the lectin domains of the F17-G, PapGII and FimH fimbrial adhesins all share the immunoglobulin-like fold of the structural components (pilins) of their fimbriae, despite lack of any sequence identity. Fold comparisons with pilin and chaperone structures of the chaperone/usher pathway highlight the central role of the C-terminal beta-strand G of the immunoglobulin-like fold and provides new insights into pilus assembly, function and adhesion., QC 20230602

The Escherichia coli adhesin F17-G is a carbohydrate-binding protein that allows the bacterium to attach to the intestinal epithelium of young ruminants. The structure of the 17 kDa lectin domain of F17-G was determined using the anomalous dispersion signal of a selenium-containing analogue of the monosaccharide ligand N-acetyl-D-glucosamine in which the anomeric oxygen was replaced by an Se atom. A three-wavelength MAD data set yielded good experimental phases to 2.6 Angstrom resolution. The structure was refined to 1.75 Angstrom resolution and was used to solve the structures of the ligand-free protein and the F17-G-N-acetyl-D-glucosamine complex. This selenium-carbohydrate phasing method could be of general use for determining the structures of carbohydrate-binding proteins., QC 20230602

The focus of this work is the synthesis of bidesmosidic saponin mimetics. Therefore, dihydrodiosgenin derivatives, which differ from the natural compounds by reduction of the 22-(hemi) acetal were used as glycosyl acceptors. In preliminary studies, the dihydrodiosgenin glycosides 16, 17 and 19, as well as trisaccharide 22, were synthesized. The acceptors 10 and 14 were subjected to DMTST-mediated glucosylation for the synthesis of these-substituted compound 3. For a selective 2,4-di-rhamnosylation of the dihydrodiosgenin glucopyranoside, differentiation of the glucose OH groups was achieved by selective benzoylation with 1-(benzoyloxy)benzotriazole. Reaction of the 3,6-di-O-benzoate 32 with the perbenzoylated ethyl thiorhamnopyranoside donor 15 gave the 2,4-di-rhamnosylated compound 33, together with the mono-rhamnosylated derivative. (, QC 20230602

Cells of the mucosal lining are the first to encounter invading bacteria during infection, and as such, they have developed numerous ways of detecting microbial intruders. Recently, we showed that epithelial cells recognize lipopolysaccharide (LPS) through the CD14-Toll-like receptor (TLR)-4 complex. Here, we identify the substructures of LPS that are recognized by the TLR4 receptor complex. In contrast to lipid A, the O-antigen does not mediate an inflammatory response; rather it interferes with the lipid A recognition. An Escherichia coli strain genetically modified to express penta-acylated lipid A not only showed reduced immunogenicity, but was also found to inhibit proinflammatory signalling induced by wild-type E. coli (hexa-acylated lipid A) as well as LPS from other bacteria of the Enterobacteriaceae family. Furthermore, penta-acylated LPS from Pseudomonas aeruginosa acted as an antagonist to hexa-acylated E. coli LPS, as did E. coli, as shown by its inhibitory effect on IL-8 production in stimulated cells. Hypo-acylated lipidA, such as that of P. aeruginosa, is found in several species within the gut microflora as well as in several bacteria causing chronic infections. Thus, our results suggest that the composition of the microflora may be important in modulating pro-inflammatory signalling in epithelial cells under normal as well as pathologic conditions.