Chemical Synthesis of Oligosaccharides Derived from Streptococcus Pneumoniae Serotype 35B and D Provides Molecular...

The wide use of Streptococcus pneumoniae capsular polysaccharide (CPS) conjugate vaccines is causing serotype replacement, and the emergence of serotype 35B is concerning because of multidrug resistance. CPS of 35B is composed of pentasaccharide repeating units that are linked through phosphodiester linkages. One of the galactofuranose residues of the pentasaccharide is acetylated, which distinguishes it from invasive serotype 35D, lacking the acetyl ester. Here, we describe a synthetic approach that can provide oligosaccharides derived of CPS 35B and 35D composed of up to 15 monosaccharides using a pentasaccharide building block equipped with four orthogonal protecting groups. The synthetic compounds were used to examine binding properties of l-ficolin, which is a protein that can activate the lectin pathway of the complement system. Solution-phase NMR experiments and computational modeling demonstrate that Galf(OAc)-1,1-Ribitol of the repeating unit of 35B CPS constitutes the minimal motif for binding to l-ficolin, and the acetyl ester is a key recognition motif. Microarray binding experiments confirmed that O-acetylation is essential for recognition and that oligosaccharides composed of 2 or 3 repeating units bind avidly due to ficolin's multimeric structures. The data provide a rationale why 35D may escape immune detection and be more invasive. The oligosaccharides were employed to investigate binding to pneumococcal serum factors 35a and 29b, which indicates that immunization with 35B CPS will not provide protection against 35D. Antibodies that can bind 35D can, however, recognize 35B, and thus, 35D CPS may provide cross-protection.