globotriaosylceramide and globotriose

Human blood contains a big variety of natural antibodies, circulating throughout life at constant concentration. Previously, we have found natural antibodies capable of binding to trisaccharide Galα1-4Galβ1-4Glc (P. To explain the inertness of human anti-P. We used a combination of immunochemical and molecular dynamics (MD) experiments. Antibodies were isolated using affinity media with P. Isolated specific antibodies were completely unable to bind natural Gb3Cer both inserted into cells and in artificial membrane, whereas strong interaction took place with synthetic analogue differing by the presence of a spacer between trisaccharide and lipid part. MD simulations revealed: i) although membrane-bound glycans do not form stable long-living aggregates, their transient packing is more compact in natural Gb3 as compared with the synthetic analog, ii) similar conformation of P. Both immunochemical and molecular dynamics data argue that the reason of the "tolerance" of natural anti-P

Topics: Animals; Antibodies; Cell Line; Chlorocebus aethiops; Epitopes; Erythrocyte Membrane; Glycolipids; Humans; Membranes, Artificial; Molecular Dynamics Simulation; Trihexosylceramides; Trisaccharides; Vero Cells

Compared to monovalent carbohydrates, multivalent carbohydrate ligands exhibit significantly enhanced binding affinities to their interacting proteins. Here, we report globotriose (P(k) ligand)-functionalized gold nanoparticle (AuNP) probes for the investigation of multivalent interactions with the B(5) subunit of Shiga-like toxin I (B-Slt). Six P(k)-ligand-encapsulated AuNPs (P(k)-AuNPs) of varying particle size and linker length were synthesized and evaluated for their potential as multivalent affinity probes by using a surface plasmon resonance competition assay. The affinity of these probes for the interacting proteins was greatly affected by nanoparticle size, linker length, and ligand density on nanoparticle surface. For example, the 20-nm 20-P(k)-l-AuNP, which had a relatively long linker showed a >10(8)-fold increase in affinity compared with the mono P(k) ligand. This intrinsic high-affinity AuNP probe specifically captured the recombinant B-Slt from Escherichia coli lysate, and the resulting purity of the B-Slt was >95 %. We also developed a robust P(k)-AuNP-based detection method for Slt-I by combining the technique with silver enhancement.

Topics: Bacteria; Biosensing Techniques; Glycoconjugates; Gold; Ligands; Metal Nanoparticles; Protein Binding; Shiga Toxin 1; Solubility; Surface Plasmon Resonance; Trihexosylceramides; Trisaccharides; Water

Shiga toxin (Stx) is a major virulence factor in infection with Stx-producing Escherichia coli (STEC). We developed a series of linear polymers of acrylamide, each with a different density of trisaccharide of globotriaosylceramide (Gb3), which is a receptor for Stx, and identified Gb3 polymers with highly clustered trisaccharides as Stx adsorbents functioning in the gut. The Gb3 polymers specifically bound to both Stx1 and Stx2 with high affinity and markedly inhibited the cytotoxic activities of these toxins. Oral administration of the Gb3 polymers protected mice after administration of a fatal dose of E. coli O157:H7, even when the polymers were administered after the infection had been established. In these mice, the serum level of Stx was markedly reduced and fatal brain damage was substantially suppressed, which suggests that the Gb3 polymers entrap Stx in the gut and prevent its entrance into the circulation. These results indicate that the Gb3 polymers can be used as oral therapeutic agents that function in the gut against STEC infections.

Topics: Acrylamide; Animals; Brain Chemistry; Carbohydrate Sequence; Disease Models, Animal; Dose-Response Relationship, Drug; Escherichia coli Infections; Escherichia coli O157; Female; Hemolytic-Uremic Syndrome; Mice; Mice, Inbred C57BL; Molecular Sequence Data; Polymers; Protein Binding; Receptors, Cell Surface; Shiga Toxin 1; Shiga Toxin 2; Shiga Toxins; Trihexosylceramides; Trisaccharides