Abstract
The coaggregation receptor polysaccharides (RPS) of Streptococcus oralis and related species are recognized by lectin-like adhesins on other members of the oral biofilm community and by RPS-specific antibodies. The former interactions involve beta-GalNAc or beta-Gal containing host-like motifs in the oligosaccharide repeating units of these polysaccharides, whereas the latter involves features of these molecules that are immunogenic. In the present investigation, the molecular and corresponding structural basis for the serotype specificity of S. oralis ATCC 10557 RPS was determined by engineering the production of this polysaccharide in transformable Streptococcus gordonii 38. This involved the systematic replacement of genes in the rps cluster of strain 38 with different but related genes from S. oralis 10557 and structural characterization of the resulting polysaccharides. The results identify four unique genes in the rps cluster of strain 10557. These include wefI for an alpha-Gal transferase, wefJ for a GalNAc-1-phosphotransferase that has a unique acceptor specificity, wefK for an acetyl transferase that acts at two positions in the hexasaccharide repeating unit, and a novel wzy associated with the beta1-3 linkage between these units. The serotype specificity of engineered polysaccharides correlated with the wefI-dependent presence of alpha-Gal in these molecules rather than with partial O-acetylation or with the linkage between repeating units. The findings illustrate a direct approach for defining the molecular basis of polysaccharide structure and antigenicity.
Highlights
The coaggregation receptor polysaccharides (RPS) of Streptococcus oralis and related species are recognized by lectin-like adhesins on other members of the oral biofilm community and by RPS-specific antibodies
We describe the rps cluster of S. oralis 10557 and the utilization of selected genes from this strain to engineer the production of type 3G RPS in transformable S. gordonii 38
Identification and Molecular Comparison of RPS Gene Clusters—The rps cluster of S. oralis 10557 (Fig. 2), which resembles those of S. oralis J22 [17] and S. oralis 34 [18], was identified downstream of dexB and two aliB-like ORFs and upstream of aliA
Summary
Firmly establishing the donor specificity of each encoded transferase (Fig. 1). In other studies, deletion of wefB eliminated L-Rha branches, converting types 2Gn and 2G RPS to linear types 1Gn and 1G, respectively [18]. Further results gained from studies of genetic complementation distinguished the GalNAc-1-phosphotransferases encoded by downstream wefC of S. gordonii 38 and wefH of S. oralis 34 by a subtle difference in acceptor specificity Both transferases acted on the linear acceptor formed in the absence of wefB, but only WefC acted on the branched acceptor formed in the presence of wefB. Genetic engineering of RPS gene clusters and structural characterization of the resulting polysaccharides provides an approach for distinguishing closely related genes such as wefH, wefC, and wefF by differences in either the donor or the acceptor specificities of their encoded transferases. These findings, which complete the comparative molecular characterization of a major RPS group [8], provide the necessary basis for tracing the evolution of these polysaccharides as recognition molecules for biofilm formation in the host oral environment
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