scholarly journals Dynamic interaction of fluoroquinolones with magnesium ions monitored using bacterial outer membrane nanopores

2020 ◽  
Vol 11 (38) ◽  
pp. 10344-10353
Author(s):  
Jiajun Wang ◽  
Jigneshkumar Dahyabhai Prajapati ◽  
Ulrich Kleinekathöfer ◽  
Mathias Winterhalter
Keyword(s):  
Outer Membrane ◽  
Divalent Cations ◽  
Dynamic Interaction ◽  
Gram Negative Bacteria ◽  
Magnesium Ions ◽  
Gram Negative ◽  
Bacterial Outer Membrane

Divalent cations alter the translocation of antibiotic molecules through the Gram-negative bacteria outer membrane nanopores.

Epitope mapping of monoclonal antibodies specific for the directly cross-linkedmeso-diaminopimelic acid peptidoglycan found in the anaerobic beer spoilage bacteriumPectinatus cerevisiiphilus

1999 ◽  
Vol 45 (9) ◽  
pp. 779-785 ◽  
Author(s):  
Barry Ziola ◽  
Sheryl L Gares ◽  
Brandene Lorrain ◽  
Lori Gee ◽  
W M Ingledew ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Outer Membrane ◽  
Epitope Mapping ◽  
Sodium Dodecyl ◽  
Diaminopimelic Acid ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Spoilage Bacteria ◽  
Beer Spoilage ◽  
Bacterial Outer Membrane

Nineteen monoclonal antibodies (Mabs) were isolated based on reactivity with disrupted Pectinatus cerevisiiphilus cells. All of the Mabs reacted with cells from which the outer membrane had been stripped by incubation with sodium dodecyl sulphate, suggesting the peptidoglycan (PG) layer was involved in binding. Mab reactivity with purified PG confirmed this. Epitope mapping revealed the Mabs in total recognize four binding sites on the PG. Mabs specific for each of the four sites also bound strongly to disrupted Pectinatus frisingensis, Selenomonas lacticifix, Zymophilus paucivorans, and Zymophilus raffinosivorans cells, but weakly to disrupted Megasphaera cerevisiae cells. No antibody reactivity was seen with disrupted cells of 11 other species of Gram-negative bacteria. These results confirm that a common PG structure is used by several species of anaerobic Gram-negative beer spoilage bacteria. These results also indicate that PG-specific Mabs can be used to rapidly detect a range of anaerobic Gram-negative beer spoilage bacteria, provided the bacterial outer membrane is first removed to allow antibody binding.Key words: beer spoilage, epitope mapping, monoclonal antibodies, Pectinatus, peptidoglycan.

scholarly journals Pal stabilises the bacterial outer membrane during constriction by a mobilisation-and-capture mechanism

2019 ◽  
Author(s):  
Joanna Szczepaniak ◽  
Peter Holmes ◽  
Karthik Rajasekar ◽  
Renata Kaminska ◽  
Firdaus Samsudin ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Outer Membrane Lipoprotein ◽  
Capture Mechanism ◽  
Dividing Cells ◽  
Bacterial Outer Membrane ◽  
Membrane Lipoprotein ◽  
Inner Membrane Protein

SummaryCoordination of outer membrane constriction with septation is critical to faithful division in Gram-negative bacteria and vital to the barrier function of the membrane. Recent studies suggest this coordination is through the active accumulation of the peptidoglycan-binding outer membrane lipoprotein Pal at division sites by the Tol system, but the mechanism is unknown. Here, we show that Pal accumulation at Escherichia coli division sites is a consequence of three key functions of the Tol system. First, Tol mobilises Pal molecules in dividing cells, which otherwise diffuse very slowly due to their binding of the cell wall. Second, Tol actively captures mobilised Pal molecules and deposits them at the division septum. Third, the active capture mechanism is analogous to that used by the inner membrane protein TonB to dislodge the plug domains of outer membrane TonB-dependent nutrient transporters. We conclude that outer membrane constriction is coordinated with cell division by active mobilisation-and-capture of Pal at division septa by the Tol system.

scholarly journals Lack of Interaction of Fluoroquinolones with Lipopolysaccharides

2002 ◽  
Vol 46 (5) ◽  
pp. 1568-1570 ◽  
Author(s):  
B. Lindner ◽  
A. Wiese ◽  
K. Brandenburg ◽  
U. Seydel ◽  
A. Dalhoff
Keyword(s):  
Outer Membrane ◽  
Antibacterial Effect ◽  
Divalent Cations ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Biologically Relevant ◽  
Mutant Strains ◽  
Trivalent Cations ◽  
Biophysical Techniques ◽  
Rough Mutant

ABSTRACT Fluoroquinolones are known to chelate with di- and trivalent cations, and it has accordingly been claimed that they perturb the integrity of the outer membrane (OM) of gram-negative bacteria. So far, chelation has not been assessed in biologically relevant test systems. Therefore, we investigated the interaction of ciprofloxacin and moxifloxacin in the absence and presence of Mg2+ with whole bacteria and isolated lipopolysaccharide (LPS) from various rough mutant strains of Salmonella enterica chemotypes by applying different biophysical techniques. We found that the fluoroquinolones did not disturb the integrity of the OM and neither were incorporated into LPS monolayers nor displaced Ca2+ from LPS monolayers, suggesting that chelation of fluoroquinolones with divalent cations does not contribute to the antibacterial effect of fluoroquinolones.

scholarly journals Bacterial Outer Membrane Vesicles as a Versatile Tool in Vaccine Research and the Fight against Antimicrobial Resistance

mBio ◽  
2021 ◽  
Author(s):  
Zhuang Zhu ◽  
Fabio Antenucci ◽  
Kasper Rømer Villumsen ◽  
Anders Miki Bojesen
Keyword(s):  
Antimicrobial Resistance ◽  
Outer Membrane ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Gram Negative Bacteria ◽  
Vaccine Research ◽  
Gram Negative ◽  
Versatile Tool ◽  
Bacterial Outer Membrane ◽  
Considerable Range

Gram-negative bacteria include a number of pathogens that cause disease in humans and animals. Although antibiotics are still effective in treating a considerable range of infections caused by Gram-negative bacteria, the alarming increase of antimicrobial resistance (AMR) induced by excessive use of antibiotics has raised global concerns.

scholarly journals Detection of Gram-negative bacterial outer membrane vesicles using DNA aptamers

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Hye-Su Shin ◽  
Vinayakumar Gedi ◽  
Joon-Ki Kim ◽  
Dong-ki Lee
Keyword(s):  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Severe Illness ◽  
Dna Aptamers ◽  
Gram Negative Bacteria ◽  
Human Beings ◽  
Gram Negative ◽  
Bacterial Outer Membrane

Abstract Infection of various pathogenic bacteria causes severe illness to human beings. Despite the research advances, current identification tools still exhibit limitations in detecting Gram-negative bacteria with high accuracy. In this study, we isolated single-stranded DNA aptamers against multiple Gram-negative bacterial species using Toggle-cell-SELEX (systemic evolution of ligands by exponential enrichment) and constructed an aptamer-based detection tool towards bacterial secretory cargo released from outer membranes of Gram-negative bacteria. Three Gram-negative bacteria, Escherichia coli DH5α, E. coli K12, and Serratia marcescens, were sequentially incubated with the pool of random DNA sequences at each SELEX loop. Two aptamers selected, GN6 and GN12, were 4.2-times and 3.6-times higher binding to 108 cells of Gram-negative bacteria than to Gram-positive bacteria tested, respectively. Using GN6 aptamer, we constructed an Enzyme-linked aptamer assay (ELAA) to detect bacterial outer membrane vesicles (OMVs) of Gram-negative bacteria, which contain several outer membrane proteins with potent immunostimulatory effects. The GN6-ELAA showed high sensitivity to detect as low as 25 ng/mL bacterial OMVs. Aptamers developed in this study show a great potential to facilitate medical diagnosis and early detection of bacterial terrorism, based on the ability to detect bacterial OMVs of multiple Gram-negative bacteria.

scholarly journals Outer Membrane Vesicle Production Facilitates LPS Remodeling and Outer Membrane Maintenance inSalmonelladuring Environmental Transitions

mBio ◽  
2016 ◽  
Vol 7 (5) ◽  
Author(s):  
Katherine E. Bonnington ◽  
Meta J. Kuehn
Keyword(s):  
Outer Membrane ◽  
Salmonella Enterica ◽  
Lipid A ◽  
Environmental Changes ◽  
Divalent Cations ◽  
Membrane Vesicle ◽  
Heterogeneous Structure ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Environmental Transitions

ABSTRACTThe ability of Gram-negative bacteria to carefully modulate outer membrane (OM) composition is essential to their survival. However, the asymmetric and heterogeneous structure of the Gram-negative OM poses unique challenges to the cell’s successful adaption to rapid environmental transitions. Although mechanisms to recycle and degrade OM phospholipid material exist, there is no known mechanism by which to remove unfavorable lipopolysaccharide (LPS) glycoforms, except slow dilution through cell growth. As all Gram-negative bacteria constitutively shed OM vesicles (OMVs), we propose that cells may utilize OMV formation as a way to selectively remove environmentally disadvantageous LPS species. We examined the native kinetics of OM composition during physiologically relevant environmental changes inSalmonella enterica, a well-characterized model system for activation of PhoP/Q and PmrA/B two-component systems (TCSs). In response to acidic pH, toxic metals, antimicrobial peptides, and lack of divalent cations, these TCSs modify the LPS lipid A and core, lengthen the O antigen, and upregulate specific OM proteins. An environmental change to PhoP/Q- and PmrA/B-activating conditions simultaneously induced the addition of modified species of LPS to the OM, downregulation of previously dominant species of LPS, greater OMV production, and increased OMV diameter. Comparison of the relative abundance of lipid A species present in the OM and the newly budded OMVs following two sets of rapid environmental shifts revealed the retention of lipid A species with modified phosphate moieties in the OM concomitant with the selective loss of palmitoylated species via vesiculation following exposure to moderately acidic environmental conditions.IMPORTANCEAll Gram-negative bacteria alter the structural composition of LPS present in their OM in response to various environmental stimuli. We developed a system to track the native dynamics of lipid A change inSalmonella entericaserovar Typhimurium following an environmental shift to PhoP/Q- and PmrA/B-inducing conditions. We show that growth conditions influence OMV production, size, and lipid A content. We further demonstrate that the lipid A content of OMVs does not fit a stochastic model of content selection, revealing the significant retention of lipid A species containing covalent modifications that mask their 1- and 4′-phosphate moieties under host-like conditions. Furthermore, palmitoylation of the lipid A to form hepta-acylated species substantially increases the likelihood of its incorporation into OMVs. These results highlight a role for the OMV response in OM remodeling and maintenance processes in Gram-negative bacteria.

Electrophysiological Characterization of Transport Across Outer Membrane Channels from Gram-Negative Bacteria in Their Native Environment

2019 ◽  
Author(s):  
Jiajun Wang ◽  
Rémi Terrasse ◽  
Jayesh Arun Bafna ◽  
Lorraine Benier ◽  
Mathias Winterhalter
Keyword(s):  
Outer Membrane ◽  
Lipid Membrane ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Multi Drug Resistance ◽  
Gram Negative Bacteria ◽  
Membrane Channels ◽  
Gram Negative ◽  
Electrophysiological Characterization

Multi-drug resistance in Gram-negative bacteria is often associated with low permeability of the outer membrane. To investigate the role of membrane channels in the uptake of antibiotics, we extract, purify and reconstitute them into artificial planar membranes. To avoid this time-consuming procedure, here we show a robust approach using fusion of native outer membrane vesicles (OMV) into planar lipid bilayer which moreover allows also to some extend the characterization of membrane protein channels in their native environment. Two major membrane channels from <i>Escherichia coli</i>, OmpF and OmpC, were overexpressed from the host and the corresponding OMVs were collected. Each OMV fusion revealed surprisingly single or only few channel activities. The asymmetry of the OMV´s translates after fusion into the lipid membrane with the LPS dominantly present at the side of OMV addition. Compared to conventional reconstitution methods, the channels fused from OMVs containing LPS have similar conductance but a much broader distribution. The addition of Enrofloxacin on the LPS side yields somewhat higher association (<i>k<sub>on</sub></i>) and lower dissociation (<i>k<sub>off</sub></i>) rates compared to LPS-free reconstitution. We conclude that using outer membrane vesicles is a fast and easy approach for functional and structural studies of membrane channels in the native membrane.

scholarly journals MexAB-OprM Efflux Pump Interaction with the Peptidoglycan of Escherichia coli and Pseudomonas aeruginosa

2021 ◽  
Vol 22 (10) ◽  
pp. 5328
Author(s):  
Miao Ma ◽  
Margaux Lustig ◽  
Michèle Salem ◽  
Dominique Mengin-Lecreulx ◽  
Gilles Phan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Cell Division ◽  
Membrane Proteins ◽  
Outer Membrane ◽  
Efflux Pump ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Active Efflux

One of the major families of membrane proteins found in prokaryote genome corresponds to the transporters. Among them, the resistance-nodulation-cell division (RND) transporters are highly studied, as being responsible for one of the most problematic mechanisms used by bacteria to resist to antibiotics, i.e., the active efflux of drugs. In Gram-negative bacteria, these proteins are inserted in the inner membrane and form a tripartite assembly with an outer membrane factor and a periplasmic linker in order to cross the two membranes to expulse molecules outside of the cell. A lot of information has been collected to understand the functional mechanism of these pumps, especially with AcrAB-TolC from Escherichia coli, but one missing piece from all the suggested models is the role of peptidoglycan in the assembly. Here, by pull-down experiments with purified peptidoglycans, we precise the MexAB-OprM interaction with the peptidoglycan from Escherichia coli and Pseudomonas aeruginosa, highlighting a role of the peptidoglycan in stabilizing the MexA-OprM complex and also differences between the two Gram-negative bacteria peptidoglycans.

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