Project Details
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Regulation of expression and biogenesis of the pneumococcal Rrg pili

Applicant Dr. Stefan Fälker
Subject Area Parasitology and Biology of Tropical Infectious Disease Pathogens
Term from 2007 to 2010
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 49967696
 
Final Report Year 2010

Final Report Abstract

Many pathogenic bacteria express and present elongated proteinaceous structures called pili on the surface to initiate contact with eukaryotic cells and other surfaces. The function and the molecular architecture of those pili have been determined in great detail in Gram-negative organisms. However, pili in Grampositive bacteria have been found much more recently and are much less well described. A fundamental difference to Gram-negative pili is the fact that those pili from Gram-positives are composed of thousands of covalently linked proteinsubunits, thereby building up a megadalton protein complex. The enzymes catalyzing this covalent linkage are called sortases, and are encoded on the chromosome in close proximity to the genes encoding the structural subunits. Within the work performed during the DFG funding period, we analyzed the biosynthesis and the regulation of the so-called Rrg pili from the important human pathogen S. pneumoniae. Together with others, we found out that those Rrg pili are composed of the main subunit RrgB in a pearl-on-a-string conformation with RrgC sitting at the proximal end and RrgA sitting at the distal end of the pilus. This is in accordance with our results clearly indicating that RrgA represent the actual adhesion of the pilus system. Therefore, this protein is especially interesting as it makes the contact with the eukaryotic cell. Furthermore, we analyzed the biosynthesis of those pili. Interestingly, we could show that only one of the three pilus-associated sortases (SrtB) is needed to build up the whole pilus, raising the question about the functional role of the other two sortases SrtC and SrtD. We could show that SrtD, together with SrtB, is needed to keep the localized wild-type presentation of the pili along the future septal zone. This localized presentation of the pili has never been described, and might have important implication for the contact between pathogen and host cell. Another important result of the project was derived from comparative structural studies of the sortase proteins. We could identify specific amino acids which are essential for catalysis and recognition of the substrate. Those results might help to identify inhibitors of sortases by rationale design, which could be new trait to fight against multi-resistant bacterial pathogens. Along this line we also identified a new DNA methyltransferase that is involved in the regulation of pilus biogenesis. Further studies are needed to estimate if this new regulatory mechanism for Gram-positives might also have the potential to be a target for antibacterial treatment. In total, the results obtained during the funding period contributed significantly to the understanding of Gram-positive pilus biology. It has become more and more obvious that the pilus systems of different Gram-positive pathogens are quite similar in the mechanism of biosynthesis and basic structure, and therefore the results for the pneumococcal Rrg pilus can give new insights also into other pilus systems. Lastly, sortases and methyltransferases might develop into new interesting targets for antibacterial treatments, which are urgently needed.

Publications

  • 2007. RrgA is a pilus-associated adhesion in Streptococcus pneumoniae. Mol Microbiol. 66:329-340
    Nelson, A. L., J. Ries, F. Bagnoli, S. Dahlberg, S. Fälker, S. Rounioja, J. Tschöp, E. Morfeldt, I. Ferlenghi, M. Hilleringmann, D. W. Holden, R. Rappuoli, S. Normark, M. A. Barocchi, & B. Henriques-Normark
  • 2008. Sortase-mediated assembly and surface topology of adhesive pneumococcal pili. Mol Microbiol. 70:595-607
    Fälker, S., A. L. Nelson, E. Morfeldt, K. Jonas, K. Hultenby, J. Ries, O. Melefors, S. Normark, & B. Henriques-Normark
  • 2009. Cloning, expression, purification, crystallization and preliminary X-ray analysis of the pilus-associated sortase C from Streptococcus pneumoniae. Acta Crystallogr Sect F Struct Biol Cryst Commun. 65:55-58
    Neiers, F., C. Madhurantakam, S. Fälker, S. Normark, B. Henriques-Normark, & A. Achour
  • 2009. Two crystal structures of pneumococcal pilus sortase C provide novel insights into catalysis and substrate specificity. J Mol Biol. 393:704-716
    Neiers, F., C. Madhurantakam, S. Fälker, C. Manzano, A. Dessen, S. Normark, B. Henriques-Normark, & A. Achour
 
 

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