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Binding of Clostridium difficile toxins to human milk oligosaccharides Open Access

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Author or creator
El-Hawiet, Amr
Kitova,Elena N.
Kitov, Pavel
Eugenio, Luiz
Ng, Kenneth K.S.
Mulvey, George L.
Dingle, Tanis C.
Szpacenko, Adam
Armstrong, Glen D.
Klassen, John S.
Additional contributors
Subject/Keyword
Mass spectrometry
Clostridium difficile
Toxin
Ligand binding
Cytotoxicity
Human milk oligosaccharide
Docking
Type of item
Journal Article (Published)
Language
English
Place
Time
Description
The binding of recombinant fragments of the C-terminal cell-binding domains of the two large exotoxins, toxin A (TcdA) and toxin B (TcdB), expressed by Clostridium difficile and a library consisting of the most abundant neutral and acidic human milk oligosaccharides (HMOs) was examined quantitatively at 25°C and pH 7 using the direct electrospray ionization mass spectrometry (ES-MS) assay. The results of the ES-MS measurements indicate that both toxin fragments investigated, TcdB-B1 and TcdA-A2, which possess one and two carbohydrate binding sites, respectively, bind specifically to HMOs ranging in size from tri- to heptasaccharides. Notably, five of the HMOs tested bind to both toxins: Fuc(α1-2)Gal(β1-4)Glc, Gal(β1-3)GlcNAc(β1-3)Gal(β1-4)Glc, Fuc(α1-2)Gal(β1-3)GlcNAc(β1-3)Gal(β1-4)Glc, Gal(β1-3)[Fuc(α1-4)]GlcNAc(β1-3)Gal(β1-4)Glc and Gal(β1-4)[Fuc(α1-3)]GlcNAc(β1-3)Gal(β1-4)Glc. However, the binding of the HMOs is uniformly weak, with apparent affinities ≤103 M−1. The results of molecular docking simulations, taken together with the experimental binding data, suggest that a disaccharide moiety (lactose or lactosamine) represents the core HMO recognition element for both toxin fragments. The results of a Verocytotoxicity neutralization assay reveal that HMOs do not significantly inhibit the cytotoxic effects of TcdA or TcdB. The absence of protection is attributed to the very weak intrinsic affinities that the toxins exhibit towards the HMOs.
Date created
2011
DOI
doi:10.7939/R3ZS2KS0K
License information
© 2011 El-Hawiet, A., Kitova, E. N., Kitov, P. I., Eugenio, L., Ng, K. K., Mulvey, G. L., Dingle, T. C., Szacenko, A., Armstrong, G. D., & Klassen, J. S. This version of this article is open access and can be downloaded and shared. The original author(s) and source must be cited.
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Citation for previous publication
El-Hawiet, A., Kitova, E. N., Kitov, P. I., Eugenio, L., Ng, K. K., Mulvey, G. L., Dingle, T. C., Szacenko, A., Armstrong, G. D., & Klassen, J. S. (2011). Binding of Clostridium difficile toxins to human milk oligosaccharides. Glycobiology, 21(9), 1217-1227.  http://doi.org/10.1093/glycob/cwr055

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File title: Binding of Clostridium difficile Toxins to Human Milk Oligosaccharides
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