Chemical structures of Streptococcus pneumoniae capsular polysaccharide type 39 (CPS39), CPS47F, and CPS34 characterized by nuclear magnetic resonance spectroscopy and their relation to CPS10A

Abstract

Structural characterization of Streptococcus pneumoniae capsular polysaccharides (CPS) is a prerequisite for unraveling both antigenic and genetic relationships that exist between different serotypes. In the current study, comparative structural studies of S. pneumoniae CPS serogroup 10 (CPS10) were extended to include genetically related S. pneumoniae CPS34, CPS39, and CPS47F. High-resolution heteronuclear nuclear magnetic resonance (NMR) spectroscopy confirmed the published structure of CPS34 and, in conjunction with glycosyl composition analyses, revealed the following repeat unit structures of the other serotypes, which have not been previously characterized: [structure: see text] Common and unique structural features of these polysaccharides, including different positions of O-acetylation, were unambiguously associated with specific genes in each corresponding cps locus. The only exception involved the gene designated wcrC, which is associated with the α1-2 transfer of Gal pyranoside (Galp) to ribitol-5-phosphate in the synthesis of CPS10A, CPS47F, and CPS34 but with α1-1 transfer of Gal to ribitol-5-phosphate in the synthesis of CPS39. The corresponding gene in the cps39 locus, although related to wcrC, more closely resembled a previously identified gene (i.e., wefM) of Streptococcus oralis that is associated with α1-1 transfer of Galp to ribitol-5-phosphate. These and other recent findings identify linkages from α-Galp to ribitol-5-phosphate and from this residue to adjacent Gal furanoside (Galf) as important sites of CPS structural and genetic diversity.

FIG 1

Central region of the multiplicity-edited ¹H-¹³C HSQC spectra of native CPS39 (A) and de-O-acetylated CPS39 (B). Negative contours (red) indicate methylene groups.

FIG 2

Structures and HMBC interresidue connectivities of CPS39 (A), CPS47F (B), and CPS34 (C).

FIG 3

Multiplicity-edited ¹H-¹³C HSQC spectra of native CPS47F (A) and de-O-acetylated CPS47F (B). Negative contours (red) indicate methylene groups. The peaks marked Y are from minor contaminating polysaccharides.

FIG 4

Genetic basis of S. pneumoniae serotype 10A, 39, 47F, and 34 CPS structures. (A) Structure-determining regions of cps loci (GenBank accession numbers CR931649, CR931711, CR931721, and CR931703, respectively), showing genes for glycosyl or ribitol-phosphate transferases (blue), polymerases (black), flippases (white), O-acetyltransferases (purple), and galactofurannose mutase (green); wciE (blue bricks) occurs as a pseudogene in the cps39 locus. (B) Association of encoded transferases and polymerases (Wzy homology group 1, 2, or 31) with each corresponding CPS structure. Gene names are the same as in previous studies (5, 6) except for wcrC in the cps39 locus, which has been renamed wefM.