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A PQS-Cleaving Quorum Quenching Enzyme Targets Extracellular Membrane Vesicles of Pseudomonas aeruginosa

Alba Arranz San Martín1, Steffen Lorenz Drees1, Susanne Fetzner1

  • 1Institute of Molecular Microbiology and Biotechnology, University of Münster, Corrensstraße 3, 48149 Münster, Germany.

Biomolecules
|November 11, 2022
PubMed

Insights

Pseudomonas aeruginosa virulence relies on quorum sensing signals. A specific enzyme, HQDsb, binds to bacterial outer membrane vesicles, enhancing the degradation of these signals.

Area of Science:

  • Microbiology and Molecular Biology
  • Bacterial Pathogenesis and Virulence
  • Enzyme Function and Regulation

Background:

  • Pseudomonas aeruginosa, an opportunistic pathogen, employs quorum sensing (QS) to regulate virulence factors.
  • The Pseudomonas quinolone signal (PQS) is a key QS molecule with membrane affinity, primarily transported via outer membrane vesicles (OMVs).
  • 3-hydroxy-4(1H)-quinolone 2,4-dioxygenases (HQDs) are known quorum quenching enzymes that cleave PQS.

Purpose of the Study:

  • To investigate the unexpected stabilization of Streptomyces bingchenggensis HQD (HQDsb) by P. aeruginosa culture supernatants.
  • To elucidate the mechanism behind HQDsb stabilization and its interaction with P. aeruginosa components.
  • To determine if HQDsb physically interacts with P. aeruginosa outer membrane vesicles (OMVs) for enhanced PQS degradation.

Main Methods:

  • Comparative analysis of HQDsb activity in culture supernatants from different P. aeruginosa strains (PA14 vs. PAO1).
  • Biochemical assays including heat treatment, lyophilization, and supernatant fractionation to identify stabilizing factors.
  • Pull-down assays to detect protein interactions between HQDsb and P. aeruginosa proteins.
  • Homo-FRET experiments to assess the spatial localization and clustering of fluorescently labeled HQDsb on prepared OMVs.

Main Results:

  • HQDsb activity was unexpectedly stabilized by P. aeruginosa culture supernatants, with a more pronounced effect from strain PA14.
  • Stabilization was independent of heat treatment and lyophilization, and small molecules were excluded as the stabilizing agents.
  • HQDsb showed interaction with P. aeruginosa proteins identified in the OMV proteome.
  • Homo-FRET confirmed the spatial clustering and binding of HQDsb to P. aeruginosa OMVs.

Conclusions:

  • The binding of HQDsb to P. aeruginosa OMVs enhances the degradation of the quorum sensing signal PQS.
  • This interaction supports the role of HQDsb as an effective quorum quenching enzyme by localizing it to the site of PQS signaling.
  • The findings provide a novel mechanism for enzyme-vesicle interaction in bacterial communication and virulence regulation.
Keywords:
Pseudomonas aeruginosaPseudomonas quinolone signaldioxygenasehomo-FRETmembrane-protein interactionouter membrane vesiclesquorum quenching enzyme

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