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Updated: May 18, 2026
Detection and Quantification of Mono-Rhamnolipids and Di-Rhamnolipids Produced by Pseudomonas aeruginosa
Published on: March 29, 2024
Functional and structural characterization of PaeM, a colicin M-like bacteriocin produced by Pseudomonas aeruginosa
Hélène Barreteau1, Mounira Tiouajni, Marc Graille
1Université Paris-Sud, Institut de Biochimie et Biophysique Moléculaire et Cellulaire, UMR 8619, F-91405 Orsay, France.
Abstract:
Colicin M (ColM) is the only enzymatic colicin reported to date that inhibits cell wall peptidoglycan biosynthesis. It catalyzes the specific degradation of the lipid intermediates involved in this pathway, thereby provoking lysis of susceptible Escherichia coli cells. A gene encoding a homologue of ColM was detected within the exoU-containing genomic island A carried by certain pathogenic Pseudomonas aeruginosa strains. This bacteriocin (pyocin) that we have named PaeM was crystallized, and its structure with and without an Mg(2+) ion bound was solved. In parallel, site-directed mutagenesis of conserved PaeM residues from the C-terminal domain was performed, confirming their essentiality for the protein activity both in vitro (lipid II-degrading activity) and in vivo (cytotoxicity against a susceptible P. aeruginosa strain). Although PaeM is structurally similar to ColM, the conformation of their active sites differs radically; in PaeM, residues essential for enzymatic activity and cytotoxicity converge toward a same pocket, whereas in ColM they are spread along a particularly elongated active site. We have also isolated a minimal domain corresponding to the C-terminal half of the PaeM protein and exhibiting a 70-fold higher enzymatic activity as compared with the full-length protein. This isolated domain of the PaeM bacteriocin was further shown to kill E. coli cells when addressed to the periplasm of these bacteria.
Insights
Researchers discovered PaeM, a Pseudomonas aeruginosa bacteriocin similar to Colicin M, which degrades lipid intermediates in cell wall biosynthesis. A minimal PaeM domain showed significantly higher enzymatic activity and killed E. coli cells.
Area of Science:
- Microbiology
- Structural Biology
- Biochemistry
Background:
- Colicin M (ColM) is a unique enzyme inhibiting bacterial cell wall peptidoglycan biosynthesis by degrading lipid intermediates.
- Pathogenic Pseudomonas aeruginosa strains carry a genomic island with a ColM homologue, termed PaeM.
Purpose of the Study:
- To characterize the structure and function of PaeM, a novel bacteriocin from Pseudomonas aeruginosa.
- To compare the enzymatic activity and active site structure of PaeM with Colicin M.
Main Methods:
- Crystallization and X-ray diffraction to solve the structure of PaeM.
- Site-directed mutagenesis to identify essential residues for PaeM activity.
- In vitro enzymatic assays measuring lipid II degradation.
- In vivo cytotoxicity assays against susceptible bacteria.
Main Results:
- PaeM structure was determined, revealing structural similarity but distinct active site conformation compared to ColM.
- Mutagenesis confirmed essential residues for PaeM's in vitro and in vivo activity.
- A minimal C-terminal domain of PaeM exhibited 70-fold higher enzymatic activity than the full-length protein.
- The isolated PaeM domain demonstrated cytotoxicity against E. coli.
Conclusions:
- PaeM represents a novel class of bacteriocins with a unique active site mechanism for inhibiting peptidoglycan biosynthesis.
- The C-terminal domain of PaeM is crucial for its enzymatic function and possesses enhanced activity.
- PaeM and its domains are potential candidates for antimicrobial development.
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