Molecular conformation affects the interaction of the Pseudomonas quinolone signal with the bacterial outer membrane

Abstract

Gram-negative bacteria produce outer-membrane vesicles (OMVs) that package genetic elements, virulence factors, and cell-to-cell communication signaling compounds. Despite their importance in many disease-related processes, how these versatile structures are formed is incompletely understood. A selfproduced secreted small molecule, the Pseudomonas quinolone signal (PQS), has been shown to initiate OMV formation in Pseudomonas aeruginosa by interacting with the outer membrane (OM) and inducing its curvature. Other bacterial species have also been shown to respond to PQS, supporting a common biophysical mechanism. Here, we conducted molecular dynamics simulations to elucidate the specific interactions between PQS and a model P. aeruginosa OM at the atomistic scale. We discovered two characteristic states of PQS interacting with the biologically relevant membrane, namely attachment to the membrane surface and insertion into the lipid A leaflet. The hydrogen bonds between PQS and the lipid A phosphates drove the PQSmembrane association. An analysis of PQS trajectory and molecular conformation revealed sequential events critical for spontaneous insertion, including probing, docking, folding, and insertion. Remarkably, PQS bent its hydrophobic side chain into a closed conformation to lower the energy barrier for penetration through the hydrophilic headgroup zone of the lipid A leaflet, which was confirmed by the potential of mean force (PMF) measurements. Attachment and insertion were simultaneously observed in the simulation with multiple PQS molecules. Our findings uncover a sequence of molecular interactions that drive PQS insertion into the bacterial OM and provide important insight into the biophysical mechanism of small molecule-induced OMV biogenesis.Outer-membrane vesicles (OMVs) 2 derived from the bacterial outer membrane (OM) are instrumental in bacterial interactions.They are spherical membranous structures that can package cargoes, such as virulence factors, proteins, DNA, RNA, and signaling molecules (1-3). In contrast to the well-known functions of OMVs in trafficking systems, the biogenesis of OMVs is still under investigation. Several mechanisms for OMV formation have been proposed (3, 4). One hypothesis builds on the accumulation of chemical compounds (e.g. misfolded proteins (5) or muramyl peptides (6)) in the periplasmic space. These fragments create turgor pressure on the OM and further stimulate vesicle formation. Another proposed mechanism focuses on reduced cross-linking between the OM and peptidoglycan layer. The lack of trans-envelope proteins untethers the OM from the cell wall (7, 8), and vesiculation occurs when the OM grows faster than the peptidoglycan layer (9). Emphasizing the membrane surface, a third explanation highlights the intercalation of molecules, such as antibiotics, phospholipids, and signaling molecules, into the outer leaflet of OM (4, 10, 11). Li et al. (10) discussed that the presence of cationic antibiotics destabilizes the OM by collapsing salt bridges between lipopolysa...

Pushing beyond the Envelope: the Potential Roles of OprF in Pseudomonas aeruginosa Biofilm Formation and Pathogenicity

“…As the OM buds into a vesicle, periplasmic contents are pulled inside, and as PQS continues to accumulate in the outer leaflet, the OM pinches off to form an independent OMV (67). Recent work from the Schertzer group supports this model, showing that conditions promoting PQS production increase OMV production (68), that PQS can promote OMV formation in various species (69), and that PQS insertion induces membrane curvature in a computational model (70).…”

Pushing the envelope: LPS modifications and their consequences

“…Another excellent example of how LPS structure influences OMV formation is the interaction of the quorum sensing molecule PQS of P. aeruginosa with lipid A 98 . PQS binds to the 4′ phosphate group of LPS, inserts partially into the membrane, and induces curvature that stimulates OMV formation 98,99 . Thus, this signaling molecule stimulates its own secretion into the environment.…”

The Role of Pseudomonas aeruginosa Lipopolysaccharide in Bacterial Pathogenesis and Physiology

“…PQS is one of the molecules of the complex Pseudomonas quorum sensing circuit, which regulates P. aeruginosa group behaviours, virulence factor production, and biofilm formation. The highly hydrophobic PQS is exported to the OM, promoting its own excretion in OMVs by interacting with Lipid A acyl chains and phosphates, which induces membrane curvature [99][100][101][102]. Remodeling of lipid A in response to environmental cues also influences OMV biogenesis.…”