AB5 toxins comprise an A subunit that corrupts essential eukaryotic cell functions, and pentameric B subunits that direct target-cell uptake after binding surface glycans. Subtilase cytotoxin (SubAB) is an AB5 toxin secreted by Shiga toxigenic Escherichia coli (STEC)1, which causes serious gastrointestinal disease in humans2. SubAB causes haemolytic uraemic syndrome-like pathology in mice3 through SubA-mediated cleavage of BiP/GRP78, an essential endoplasmic reticulum chaperone4. Here we show that SubB has a strong preference for glycans terminating in the sialic acid N-glycolylneuraminic acid (Neu5Gc), a monosaccharide not synthesized in humans. Structures of SubB-Neu5Gc complexes revealed the basis for this specificity, and mutagenesis of key SubB residues abrogated in vitro glycan recognition, cell binding and cytotoxicity. SubAB specificity for Neu5Gc was confirmed using mouse tissues with a human-like deficiency of Neu5Gc and human cell lines fed with Neu5Gc. Despite lack of Neu5Gc biosynthesis in humans, assimilation of dietary Neu5Gc creates high-affinity receptors on human gut epithelia and kidney vasculature. This, and the lack of Neu5Gc-containing body fluid competitors in humans, confers susceptibility to the gastrointestinal and systemic toxicities of SubAB. Ironically, foods rich in Neu5Gc are the most common source of STEC contamination. Thus a bacterial toxin's receptor is generated by metabolic incorporation of an exogenous factor derived from food.
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Protein Data Bank
The coordinates and structure factors for the SubB structures are deposited in Protein Data Bank under accession numbers 3DWA, 3DWP and 3DWQ. Raw glycan array data are available at http://www.functionalglycomics.org/glycomics/publicdata/selectedScreens.jsp.
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We thank the staff at the General Medicine and Cancer Institutes Collaborative Access Team Advanced Photon Source, Chicago, for assistance with data collection. This research was supported by a Program Grant from the National Health and Medical Research Council of Australia (NHMRC; to A.W.P. and J.C.P.), an NHMRC Project Grant (to T.B. and A.W.P.), a grant from the National Institute of General Medical Sciences to the Consortium for Functional Glycomics, RO1 grants from the National Institutes of Health (to A.W.P., J.C.P., J.R., X.C. and A.V.) and from the ARC Centre of Excellence in Structural and Functional Microbial Genomics (to J.R.). J.R. is supported by an Australian Research Council Federation Fellowship; T.B. by an NHMRC Career Development Award; J.C.P. by an NHMRC Australia Fellowship. We also thank C. J. Gregg for assistance with in vivo experiments and with collection of data, and L. Wiggleton for technical assistance with tissue sectioning and staining.
Author Contributions E.B. and A.W.P. contributed equally. E.B, T.B. and M.C.J.W. crystallized SubB, solved the structure and contributed to manuscript preparation. A.W.P. constructed mutants and contributed to design and interpretation of experiments, project management and writing of the manuscript. J.C.P. and J.R. contributed to design and interpretation of experiments, project management and writing of the manuscript. D.C.C. and U.M.T. performed experiments. D.F.S. performed and interpreted glycan array experiments. J.C.L., N.M.V. and A.V. designed, performed and interpreted experiments relating to Neu5Gc on cells and tissues and to cytotoxicity in vivo, and contributed to manuscript preparation. H.Y., S.H. and X.C. synthesized oligosaccharides. A.V., T.B. and J.R. are joint senior and corresponding authors.
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Byres, E., Paton, A., Paton, J. et al. Incorporation of a non-human glycan mediates human susceptibility to a bacterial toxin. Nature 456, 648–652 (2008). https://doi.org/10.1038/nature07428
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