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A Purification and In Vitro Activity Assay for a pppGpp Synthetase from Clostridium difficile
Published on: November 3, 2018
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Disparate subcellular location of putative sortase substrates in Clostridium difficile
Johann Peltier1,2, Helen A Shaw1,2, Brendan W Wren2
1Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK.
Scientific Reports
|August 25, 2017
Summary
Clostridium difficile sortase SrtB anchors specific proteins to the cell wall via an SPKTG motif, crucial for pathogen colonization. This mechanism
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Cell Wall Biology
Background:
- Clostridium difficile is a significant gastrointestinal pathogen.
- Understanding C. difficile colonization mechanisms is crucial.
- Sortase enzymes are key for anchoring bacterial surface proteins.
Purpose of the Study:
- To investigate the role of sortase enzymes in C. difficile colonization.
- To identify functional sortase substrates in C. difficile.
- To elucidate the mechanism of protein anchoring to the C. difficile cell wall.
Main Methods:
- In vitro analysis of sortase SrtB activity.
- Site-directed mutagenesis of C-terminal sorting motifs.
- In vivo asymmetric cleavage assays.
Main Results:
- C. difficile proteins CD2537 and CD3392 are identified as functional SrtB substrates.
- The SPKTG motif is essential for covalent cell wall attachment.
- Proteins with similar motifs (SPSTG, SPQTG) are not anchored by sortase.
- SrtB-dependent localization is critical for protein function, even with conserved motifs.
Conclusions:
- Sortase SrtB plays a vital role in anchoring specific C. difficile proteins to the cell wall.
- The SPKTG motif is a key determinant for sortase-mediated anchoring.
- Differential substrate recognition by sortase influences protein localization and potentially pathogenicity.
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