The Beauty of Asymmetric Membranes: Reconstitution of the Outer Membrane of Gram-Negative Bacteria

Abstract

The architecture of the lipid matrix of the outer membrane of Gram-negative bacteria is extremely asymmetric: Whereas the inner leaflet is composed of a phospholipid mixture, the outer leaflet is built up by glycolipids. For most Gram-negative species, these glycolipids are lipopolysaccharides (LPS), for a few species, however, glycosphingolipids. We demonstrate experimental approaches for the reconstitution of these asymmetric membranes as (i) solid supported membranes prepared by the Langmuir-Blodgett technique, (ii) planar lipid bilayers prepared by the Montal-Mueller technique, and (iii) giant unilamellar vesicles (GUVs) prepared by the phase transfer method. The asymmetric GUVs (aGUVs) composed of LPS on one leaflet are shown for the first time. They are characterized with respect to their phase behavior, flipflop of lipids and their usability to investigate the interaction with membrane active peptides or proteins. For the antimicrobial peptide LL-32 and for the bacterial porin OmpF the specificity of the interaction with asymmetric membranes is shown. The three reconstitution systems are compared with respect to their usability to investigate domain formation and interactions with peptides and proteins.

Lipid-mediated antimicrobial resistance: a phantom menace or a new hope?

“…The transport of lipids between membranes or across membrane leaflets can mediate changes in membrane structure and biophysical properties that limit the interaction of antimicrobials with the membrane, affecting antimicrobial susceptibility (Fig. 1F ) (May and Grabowicz 2018 ; Bogdanov et al 2020 ; Paulowski et al 2020 ). In the outer membrane of Gram-negative bacteria, a number of regulatory systems help maintain the highly asymmetric distribution of lipopolysaccharides between the inner and outer leaflet, required for both membrane integrity and resistance to antimicrobials (May and Grabowicz 2018 ).…”

Molecular Mechanisms of Lipid Metabolism Disorders in Infectious Exacerbations of Chronic Obstructive Pulmonary Disease

“…Some pathogens can affect the lipid metabolism in the host cells by realizing their virulence, as well as using the host lipids as a food source or assimilating them into their metabolic or structural processes [ 280 ]. For colonization of the lungs in COPD bacteria use many other strategies, including modification of the lipid composition of the membrane, which affects its permeability for antibacterial drugs, the formation of biofilms, and the exclusion of certain lipids from the host metabolic and immune processes [ 281 ].…”

Loss of β-Ketoacyl Acyl Carrier Protein Synthase III Activity Restores Multidrug-Resistant Escherichia coli Sensitivity to Previously Ineffective Antibiotics

“…The PL-LPS ratio, maintenance of membrane asymmetry, surface charge profiles of the surface exposed LPS, and chemical differences within both the PLs and LPS components collectively define the overall physicochemical and biological properties of the OM ( 22 ). As such, progress in understanding the OM may lead to valuable tactics to circumvent this formidable protective barrier and improve antibiotic cell entry in Gram-negative bacteria.…”