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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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Two small (p)ppGpp synthases in Staphylococcus aureus mediate tolerance against cell envelope stress conditions
Tobias Geiger1, Benjamin Kästle, Fabio Lino Gratani
1Interfaculty Institute of Microbiology and Infection Medicine, University of Tübingen, Tübingen, Germany.
Journal of Bacteriology
|December 17, 2013
Summary
Staphylococcus aureus uses RelP and RelQ (p)ppGpp synthases to survive cell envelope stress. These enzymes are induced by antibiotics, aiding bacterial tolerance and survival.
Area of Science:
- Microbiology
- Bacterial Physiology
- Molecular Biology
Background:
- The stringent response, mediated by (p)ppGpp nucleotides, is a crucial bacterial survival mechanism.
- Staphylococcus aureus has three (p)ppGpp synthases: RSH, RelP, and RelQ.
Purpose of the Study:
- To characterize the biochemical activities and in vivo roles of the small (p)ppGpp synthases, RelP and RelQ, in S. aureus.
- To investigate the function of RelP and RelQ in bacterial stress response and antibiotic tolerance.
Main Methods:
- Purification and biochemical analysis of RelP and RelQ proteins.
- In vivo studies using S. aureus strains with modified gene expression.
- Transcriptional analysis of relP and relQ expression under antibiotic stress.
- Assessment of mutant survival rates upon exposure to cell wall-active antibiotics.
Main Results:
- RelP exhibits stronger synthetic activity than RelQ.
- Both RelP and RelQ prefer GDP over GTP for (p)ppGpp synthesis.
- RelP and RelQ are essential for preventing toxic accumulation of (p)ppGpp, as demonstrated by growth arrest and cell death in hydrolase-negative backgrounds.
- relP and relQ expression is significantly induced by vancomycin and ampicillin.
- Mutants lacking relP and relQ display reduced survival against cell wall-active antibiotics.
Conclusions:
- RelP and RelQ are active (p)ppGpp synthases in S. aureus.
- These enzymes are induced during cell envelope stress, contributing to antibiotic tolerance.
- RelP and RelQ play vital roles in S. aureus survival under antibiotic pressure.
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