Recognition of human milk oligosaccharides by bacterial exotoxins.
The affinities of the most abundant oligosaccharides found in human milk for four bacterial exotoxins (from Vibrio cholerae and pathogenic Escherichia coli) were quantified for the first time. Association constants (Ka) for a library of 20 human milk oligosaccharides (HMOs) binding to Shiga toxin type 2 holotoxin (Stx2) and the B subunit homopentamers of cholera toxin, heat-labile toxin and Shiga toxin type 1 (CTB5, HLTB5 and Stx1B5) were measured at 25°C and pH 7 using the direct electrospray ionization mass spectrometry assay. Notably, all four bacterial toxins bind to a majority of the HMOs tested and five of the HMOs (2'-fucosyllactose, lacto-N-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II and lacto-N-fucopentaose III) are ligands for all four toxins. These five HMOs are also reported to bind to other bacterial toxins (e.g. toxin A and toxin B of Clostridium difficile). In all cases, the HMO affinities (apparent Ka) are relatively modest (≤15,000 M(-1)). However, at the high concentrations of HMOs typically ingested by infants, a significant fraction of these toxins, if present, is expected to be bound to HMOs. Binding measurements carried out with 2'-fucosyllactose or lacto-N-fucopentaose I, together with a high-affinity ligand based on the native carbohydrate receptor, revealed that all four toxins possess HMO-binding sites that are distinct from those of the native receptors, although evidence of competitive binding was found for lacto-N-fucopentaose I with Stx2 and 2'-fucosyllactose and lacto-N-fucopentaose I with HLTB5. Taken together, the results of this study suggest that, while HMOs are expected to bind extensively to these bacterial toxins, it is unlikely that HMO binding will effectively inhibit their interactions with their cellular receptors.
Alberta Glycomics Centre and Department of Chemistry, University of Alberta, Edmonton, AB, Canada T6G 2G2.
Amr
A
El-Hawiet
Alberta Glycomics Centre and Department of Chemistry, University of Alberta, Edmonton, AB, Canada T6G 2G2.
Elena N
EN
Kitova
Alberta Glycomics Centre and Department of Chemistry, University of Alberta, Edmonton, AB, Canada T6G 2G2 john.klassen@ualberta.ca.
John S
JS
Klassen
D000606:Amino Sugars / Q000737:chemistry / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification; D001427:Bacterial Toxins / Q000737:chemistry / Q000302:isolation & purification; D001665:Binding Sites; D002240:Carbohydrate Sequence; D002772:Cholera Toxin / Q000737:chemistry / Q000302:isolation & purification; D016360:Clostridioides difficile / Q000737:chemistry*; D054308:Enteropathogenic Escherichia coli / Q000737:chemistry*; D004768:Enterotoxins / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008895:Milk, Human / Q000737:chemistry*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000302:isolation & purification; D011134:Polysaccharides / Q000737:chemistry / Q000302:isolation & purification; D011485:Protein Binding; D022622:Shiga Toxin 1 / Q000737:chemistry / Q000302:isolation & purification; D022641:Shiga Toxin 2 / Q000737:chemistry / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization; D014312:Trisaccharides / Q000737:chemistry / Q000302:isolation & purification; D014734:Vibrio cholerae / Q000737:chemistry*
D000606:Amino Sugars; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004768:Enterotoxins; D009844:Oligosaccharides; D011134:Polysaccharides; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D014312:Trisaccharides; C434666:lacto-N-fucopentaose III; C450841:shiga toxin 2 B subunit; C041348:tcdA protein, Clostridium difficile; C031815:toxB protein, Clostridium difficile; C041762:lacto-N-fucopentaose II; C045442:lacto-N-fucopentaose I; D002772:Cholera Toxin; C013084:lacto-N-neotetraose; C031420:2'-fucosyllactose
affinity; electrospray ionization mass spectrometry; human milk oligosaccharide; toxin