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M010287200v1
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Papers In Press, published online ahead of print March 22, 2001
J. Biol. Chem, 10.1074/jbc.M010287200
Submitted on November 13, 2000
Revised on March 9, 2001
Accepted on March 22, 2001

Tts, a processive beta -glucosyltransferase of Streptococcus pneumoniae, directs the synthesis of the branched type 37 capsular polysaccharide in pneumococcus and other Gram-positive species

Daniel Llull, Ernesto García, and Rubens López

Centro de Investigaciones Biologicas, Consejo Superior de Investigaciones Cientificas, 28006, Madrid

Corresponding Author: e.garcia{at}cib.csic.es

The type 37 capsule of Streptococcus pneumoniae is a homopolisaccharide built up from repeating units of [beta -D-Glcp-(1->2)]-beta -D-Glcp-(1->3). The elements governing the expression of the tts gene, coding for the glucosyltransferase involved in the synthesis of the type 37 pneumococcal capsular polysaccharide, have been studied. Primer extension analysis and functional tests demonstrated the presence of four new transcriptional start points upstream of the previously reported tts promoter (ttsp). Interestingly, three of these transcriptional start points are located in a RUP element thought to be involved in recombinational events (Oggioni, M. R., and Claverys, J. P. (1999) Microbiology 145, 2647?2653). Transformation experiments using either a recombinant plasmid containing the whole transcriptional unit of tts or chromosomal DNA from a type 37 pneumococcus showed that tts is the only gene required to drive the biosynthesis of a type 37 capsule in S. pneumoniae and other Gram-positive bacteria, namely, Streptococcus oralis, Streptococcus gordonii, and Bacillus subtilis. The Tts synthase was overproduced in S. pneumoniae and purified as a membrane-associated enzyme. These membrane preparations used UDP-Glc as substrate to catalyze the synthesis of a high molecular weight polysaccharide immunologically identical to the type 37 capsule. In addition, UDP-Gal was also a substrate to produce type 37 polysaccharide since a strong UDP-Glc-4?-epimerase activity is associated to the membrane fraction of S. pneumoniae. These results indicated that Tts has a dual biochemical activity that leads to the synthesis of the branched type 37 polysaccharide.


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