First Evidence for a Covalent Linkage between Enterobacterial Common Antigen and Lipopolysaccharide in Shigella sonnei Phase II ECALPS |
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Authors: | Tomasz K Gozdziewicz Czeslaw Lugowski Jolanta Lukasiewicz |
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Institution: | From the ‡Department of Immunochemistry, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, R. Weigla 12, PL-53-114 Wroclaw, Poland and ;the §Department of Biotechnology and Molecular Biology, University of Opole, kard. B. Kominka 6A, PL-45-035 Opole, Poland |
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Abstract: | Enterobacterial common antigen (ECA) is expressed by Gram-negative bacteria belonging to Enterobacteriaceae, including emerging drug-resistant pathogens such as Escherichia coli, Klebsiella pneumoniae, and Proteus spp. Recent studies have indicated the importance of ECA for cell envelope integrity, flagellum expression, and resistance of enteric bacteria to acetic acid and bile salts. ECA, a heteropolysaccharide built from the trisaccharide repeating unit, →3)-α-d-Fucp4NAc-(1→4)-β-d-ManpNAcA-(1→4)-α-d-GlcpNAc-(1→, occurs as a cyclic form (ECACYC), a phosphatidylglycerol (PG)-linked form (ECAPG), and an endotoxin/lipopolysaccharide (LPS)-associated form (ECALPS). Since the discovery of ECA in 1962, the structures of ECAPG and ECACYC have been completely elucidated. However, no direct evidence has been presented to support a covalent linkage between ECA and LPS; only serological indications of co-association have been reported. This is paradoxical, given that ECA was first identified based on the capacity of immunogenic ECALPS to elicit antibodies cross-reactive with enterobacteria. Using a simple isolation protocol supported by serological tracking of ECA epitopes and NMR spectroscopy and mass spectrometry, we have succeeded in the first detection, isolation, and complete structural analysis of poly- and oligosaccharides of Shigella sonnei phase II ECALPS. ECALPS consists of the core oligosaccharide substituted with one to four repeating units of ECA at the position occupied by the O-antigen in the case of smooth S. sonnei phase I. These data represent the first structural evidence for the existence of ECALPS in the half-century since it was first discovered and provide insights that could prove helpful in further structural analyses and screening of ECALPS among Enterobacteriaceae species. |
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Keywords: | Bacteria Endotoxin Lipopolysaccharide (LPS) Mass Spectrometry (MS) NMR ECA Shigella Enterobacterial Common Antigen |
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