C-Terminal detection of the Streptococcus pneumoniae GalU protein causes decrease in capsular polysaccharide production

BONOFIGLIO L; MOSCOSO NAYA M; GARCÍA E; MOLLERACH M

Simposio; 8th International Symposium on Pneumococci and Pneumococcal Diseases; 2012

ISPPD Association

The GalU protein plays a key role in polysaccharide byosinthesis. To better understand the implication of this protein in capsule production, galU gene was interrupted by mariner mutagenesis. The derivative mutant obtained of S. pneumoniae R6 presented an insertion of Himar1 minitransposon in the last 101 nucleotides of the galU gene. DNA obtained from this mutant was used to transform the encapsulated M23 strain (serotype 3). Transformants were analyzed by PCR mapping and sequencing. M23c4 mutant has a deletion of the 33 C-terminal amino acids in GalU relative to the full length protein. Agglutination with S3 antiserum was observed under phase contrast microscopy, although M23c4 has rough colony morphology. Polysaccharide extraction was performed from wild type and M23c4 mutant; semiquantitative double-diffusion experiments indicated that a less quantitative polysaccharide is produced in the mutant compared to wild type strain. A model of the truncated protein was performed using Swiss model and Deep-Viewer 4.0 using E. coli protein as template. This protein is a tetramer composed by two dimers in which the monomeric subunits are tightly intertwined. According to our model, the mutated GalU has lost the helix involved in the subunit-subunit interaction of the tight dimer, producing an altered quaternary structure of the protein. This modification could explain the decrease in polysaccharide production probably due to a lower enzymatic activity of GalU. These results contribute to the knowledge of GalU enzyme, which is considered a target for the search of new antipneumococcal drugs.