scholarly journals CpsR, a GntR family regulator, transcriptionally regulates capsular polysaccharide biosynthesis and governs bacterial virulence in Streptococcus pneumoniae

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Kaifeng Wu ◽  
Hongmei Xu ◽  
Yuqiang Zheng ◽  
Libin Wang ◽  
Xuemei Zhang ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Bacterial Virulence ◽  
Polysaccharide Biosynthesis ◽  
Gntr Family

Identification of Streptococcus pneumoniae genes specifically induced in mouse lung tissues

2008 ◽  
Vol 54 (1) ◽  
pp. 58-65 ◽  
Author(s):  
Jiang-Ping Meng ◽  
Yi-Bing Yin ◽  
Xue-Mei Zhang ◽  
Yuan-Shuai Huang ◽  
Kai Lan ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Mouse Lung ◽  
Selection System ◽  
Polysaccharide Biosynthesis ◽  
Dual Reporter ◽  
Promoter Trap ◽  
Substance Transport ◽  
Adherence Energy

To identify Streptococcus pneumoniae genes expressed specifically during infections, a selection system based on the in vivo expression technology (IVET) was established. galU, which is critical for capsular polysaccharide biosynthesis, and lacZY encoding β-galactosidase were employed as dual reporter genes to screen in-vivo-induced (ivi) genes of S. pneumoniae. The galU-deficient mutant of S. pneumoniae is incapable of utilizing galactose, thus failing to synthesize capsular polysaccharide, and therefore loses its ability to survive in the host. A promoter-trap library was constructed in S. pneumoniae, which was used to infect BALB/c mice in an intranostril model. Those strains recovered from lung tissue of mice and exhibiting a white colony phenotype on tryptic soy agar containing X-gal (5-bromo-4-chloro-3-indolyl-β-d-galactopyranoside) were collected and identificated. A total of 15 unique sequences were obtained through in vivo screening. The ivi genes of S. pneumoniae are involved in many processes, such as colonization and adherence, energy metabolism, nutrient substance transport, transcription regulation, DNA metabolism, and cell wall synthesis. There are some hypothetical proteins whose functions are not clear. This novel IVET is a useful tool for identifying ivi genes in S. pneumoniae.

scholarly journals The capsular polysaccharide biosynthesis of Streptococcus pneumoniae serotype 8: functional identification of the glycosyltransferase WciS (Cap8H)

2005 ◽  
Vol 389 (1) ◽  
pp. 63-72 ◽  
Author(s):  
Nehmé SAKSOUK ◽  
Ludovic PELOSI ◽  
Pierre COLIN-MOREL ◽  
Manel BOUMEDIENNE ◽  
Patricia L. ABDIAN ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Biochemical Characterization ◽  
Repeat Unit ◽  
Sugar Residue ◽  
Functional Identification ◽  
Polysaccharide Biosynthesis ◽  
Sugar Addition ◽  
Galactose Residue

CPS (capsular polysaccharide) is a major virulence factor in Streptococcus pneumoniae. Biosynthesis of CPS RU (repeat unit) proceeds by sequential transfer of sugar residues from the appropriate sugar donor to an activated lipid carrier by committed GTs (glycosyltransferases). While the nucleotide sequence of many cps loci is already known, the real substrate specificity of the hypothetical GTs, as well as the sequence of sugar addition is unclear. In the present paper, we report the biochemical characterization of one α-galactosyltransferase, WciS (Cap8H), a member of GT family 4. This enzyme is implicated in the tetrasaccharide RU biosynthetic pathway of Strep. pneumoniae CPS 8 ([→4)-α-D-Glcp-(1→4)-α-D-Galp-(1→4)-β-D-GlcAp-(1→4)-β-D-Glcp-(1→]n). Expression of WciS–His6 in Escherichia coli BL21 (DE3) strains or BL21 (DE3)/ΔgalU strain resulted in synthesis of a 39 kDa membrane-associated protein identified by N-terminal sequencing and recognized by anti-His6-tag antibody. This protein was capable of adding a galactose residue cellobiuronic acid [β-D-GlcAp-(1→4)-D-Glcp]-pyrophosphate-polyprenol from UDP-Gal. The newly added galactose residue is removed by α-galactosidase, indicating that WciS is a retaining GT. Our results suggest that WciS catalyses the addition of the third sugar residue of the CPS 8 RU. The recombinant WciS–His6 was solubilized and purified as a soluble multimer, opening the way for structural studies.

scholarly journals Analysis of the 5′ Portion of the Type 19A Capsule Locus Identifies Two Classes of cpsC, cpsD, andcpsE Genes in Streptococcus pneumoniae

1999 ◽  
Vol 181 (11) ◽  
pp. 3599-3605 ◽  
Author(s):  
Judy K. Morona ◽  
Renato Morona ◽  
James C. Paton
Keyword(s):  
Streptococcus Pneumoniae ◽  
Southern Hybridization ◽  
Capsular Polysaccharide ◽  
Sequence Data ◽  
Direct Sequencing ◽  
Southern Hybridization Analysis ◽  
Polysaccharide Biosynthesis ◽  
Capsule Locus ◽  
Pcr Products ◽  
Long Range Pcr

ABSTRACT Analysis of the sequence data obtained from the 5′ portion of theStreptococcus pneumoniae type 19A capsular polysaccharide biosynthesis locus (cps19a) revealed that the first seven genes are homologous to the first seven genes in the type 19F (cps19f) locus. The former genes were designatedcps19aA to -G and were 70 to 90% identical to their cps19f counterparts. Southern hybridization analysis of the cps loci from various S. pneumoniaeserotypes with probes specific for the cps19aC,cps19aD, and cps19aE genes indicated a hybridization pattern complementary to that previously reported forcps19fC, cps19fD, and cps19fE. That is, all serotypes tested contained high-stringency homologues of either the cps19aC to -E genes or thecps19fC to -E genes, but not both. On this basis S. pneumoniae cps loci can be divided into two distinct classes. Long-range PCR was used to amplify thecps regions between cpsB and aliAfrom a variety of pneumococcal serotypes. Direct sequencing of the 5′ end of these PCR products, and phylogenetic analysis of the sequence data, confirmed the presence of the two distinct classes ofcpsC. Whereas members within one class are greater than 95% identical to each other, the DNA sequence identity between the two classes is only approximately 70%.

scholarly journals Characterization of the galU Gene of Streptococcus pneumoniae Encoding a Uridine Diphosphoglucose Pyrophosphorylase: A Gene Essential for Capsular Polysaccharide Biosynthesis

1998 ◽  
Vol 188 (11) ◽  
pp. 2047-2056 ◽  
Author(s):  
Marta Mollerach ◽  
Rubens López ◽  
Ernesto García
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
New Drugs ◽  
Identical Result ◽  
Specific Gene ◽  
Polysaccharide Biosynthesis ◽  
Escherichia Coli Cells ◽  
Type 3

The galU gene of Streptococcus pneumoniae has been cloned and sequenced. Escherichia coli cells harboring the recombinant plasmid pMMG2 (galU) overproduced a protein that has been shown to correspond to a uridine 5′-triphosphate:glucose-1-phosphate uridylyltransferase (uridine diphosphoglucose [UDP-Glc] pyrophosphorylase) responsible for the synthesis of UDP-Glc, a key compound in the biosynthesis of polysaccharides. A gene very similar to the S. pneumoniae galU has been found in a partial nucleotide sequence of the Streptococcus pyogenes genome. Knockout galU mutants of type 1 pneumococci are unable to synthesize a detectable capsule. An identical result was found in type 3 S. pneumoniae cells in spite of the fact that these bacteria contain a type-specific gene (cap3C) that also encodes a UDP-Glc pyrophosphorylase. Since eukaryotic UDP-Glc pyrophosphorylases appear to be completely unrelated to their prokaryotic counterparts, we postulate that GalU may be an appropriate target for the search of new drugs to control the pathogenicity of bacteria like pneumococcus and S. pyogenes.

scholarly journals Identification of the cpsA gene as a specific marker for the discrimination of Streptococcus pneumoniae from viridans group streptococci

2010 ◽  
Vol 59 (10) ◽  
pp. 1146-1152 ◽  
Author(s):  
Hee Kuk Park ◽  
Sang-Jae Lee ◽  
Jang Won Yoon ◽  
Jong Wook Shin ◽  
Hyoung-Shik Shin ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Dna Sequence ◽  
Dna Sequences ◽  
Capsular Polysaccharide ◽  
Subtractive Hybridization ◽  
Rapid Identification ◽  
Specific Marker ◽  
Viridans Streptococci ◽  
Polysaccharide Biosynthesis ◽  
Viridans Group Streptococci

Streptococcus pneumoniae, the aetiological agent of pneumonia and non-gonococcal urethritis, shares a high degree of DNA sequence identity with the viridans group of streptococci, particularly Streptococcus mitis and Streptococcus oralis. Although their clinical and pathological manifestations are different, discrimination between S. pneumoniae and its close viridans cocci relatives is still quite difficult. Suppression subtractive hybridization was performed to identify the genomic differences between S. pneumoniae and S. mitis. Thirty-four resulting S. pneumoniae-specific clones were examined by sequence determination and comparative DNA sequence analysis using blast. S. pneumoniae-specific primers were subsequently designed from one of the clonal DNA sequences containing the cps gene (coding for capsular polysaccharide biosynthesis). The primer specificities were evaluated using 49 viridans streptococci including 26 S. pneumoniae, 54 other streptococci, 14 Lactococcus species, 14 Enterococcus species and three Vagococcus species, and compared with the specificities of previously described autolysin (lytA), pneumolysin (ply), Spn9802 and Spn9828 primers. The newly designed cpsA-specific primer set was highly specific to S. pneumoniae and was even better than the existing primers. These findings may help improve the rapid identification and differentiation of S. pneumoniae from closely related members of the viridans group streptococci.

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