The Shigella flexneri O‐antigenic polysaccharide chain Nature of the biological repeating unit

Abstract

The sequence of monosaccharides in the biological repeating tetrasaccharide unit of Shigella flexneri variant Y O-antigenic polysaccharide chain was determined by subjecting three oligosaccharides of the polysaccharide, obtained by phage-Sf6-mediated enzymatic hydrolysis, to methylation analysis and proton nuclear magnetic resonance spectroscopy. The smallest saccharide was shown to be a tetrasaccharide with the structure α-l-Rhap-(1–3)-β-d-GlcpNA_c-(1–2)-α-l-Rhap-(1–2)-l-Rha. The next saccharide, an octasaccharide, was shown to be a dimer of the tetrasaccharide with the l-Rha residues linked α1.3. The longest saccharide was shown to be a decasaccharide with the following structure: α-l-Rhap-(1–2)-α-l-Rhap-(1–3)-α-l-Rhap-(1–3)-β-d-GlcpNAc-(1–2)-α-l-Rhap-(1–2)-α-l-Rhap-(1–3)-α-l-Rhap-(1–3)-β-l-GlcpNAc-(1–2)-α-l-Rhap-(1–2)-l-Rha. Thus the decasaccharide differed from the octasaccharide and tetrasaccharide by having the α-l-Rhap-(1–2)-α-Rhap disaccharide added in the terminal non-reducing end of the saccharide chain. This shows that the α-l-Rhap-(1–2)-α-l-Rhap-(1–2)-α-l-Rhap-(1–3)-d-GlcpNAc tetrasaccharide is the biological repeating unit of the O chain and that the repeating units are joined through a β-d-GlcpNAc-(1–2)-l-Rhap linkage. Inhibition experiments utilizing the enzyme-linked immunosorbent assay (ELISA) with S. flexneri Y lipopolysaccharide/S. flexneri Y rabbit antiserum showed that the decasaccharide was the best inhibitor (threefold as active as the octasaccharide and sixtyfold as active as the tetrasaccharide); this supports the postulated structure of the biological repeating unit.