The capsular polysacchrides of Streptococcus pneumoniae represent the most important virulence factor of this organism. Ninety structurally and antigenically diverse capsules have been identified, and the synthesis of all proceeds by either the Wzy-polymerase-dependent mechanism or the synthase-dependent mechanism. Synthesis of the type 3 capsule [-4)-beta-D-Glc-(1-3)-beta-D-GlcUA-(1-] is representative of the synthase-dependent mechanism, and its analysis serves as a paradigm for polysaccharide synthesis by processive beta-glycosyltransferases in both prokaryotic and eukaryotic organisms. Among the related enzymes in the glycosyltransferase 2 family are the chitin, cellulose, and hyaluronan synthases. Analysis of type 3 synthesis has identified novel aspects of the biosynthetic mechanism that may be broadly applicable to polymers in this family. Synthesis of the type 3 polymer initiates by the transfer of glucose (Glc) to phosphatidylglycerol and proceeds by the alternate addition of glucuronic acid (GlcUA) and Glc to the non-reducing end of the growing polymer. With sufficient UDP-GIc and UDP-GlcUA, the oligosaccharide-lipid reaches the length of an octasaccharide, at which point it becomes tightly bound to the carbohydrate-binding site of the synthase, and a highly processive reaction leading to high molecular weight polymer ensues. The transition from oligosaccharide-lipid to polysaccharide synthesis is enhanced by increasing UDP-GlcUA concentrations and reduced by limiting UDP-GlcUA. Both in vitro and in vivo, the length of the polysaccharide chain is determined by the concentration of UDP-GlcUA under high UDP-GIc concentrations, as occurs in the cell. Chain termination results in ejection of the polymer from the synthase and, if the linkage to PG has been disrupted, polymer release from the membrane and from the cell. These results have led to models for the control of initiation, synthesis, and chain length determination for type 3 and related polymers. Further analysis of type 3 synthesis will allow us to characterize: 1) reactions mediated by the type 3 synthase, 2) factors that influence synthase activity, 3) mechanisms that control cellular UDP-GlcUA concentrations and capsule chain length, and 4) mechanisms involved in release of capsule from the cell. The relevance of these properties to the pathogenesis of S. pneumoniae will also be assessed, allowing us to better understand an essential virulence factor of this and many other bacteria.
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