Correlative microscopy reveals the nanoscale morphology of E. coli-derived supported lipid bilayers

Supported lipid bilayers (SLBs) made from reconstituted lipid vesicles are an important tool in molecular biology. A breakthrough in the field has come with the use of vesicles derived from cell membranes to form SLBs. These new supported bilayers, consisting both of natural and synthetic components, provide a physiologically relevant system on which to study protein-protein interactions as well as protein-ligand interactions and other lipid membrane properties. These complex bilayer systems hold promise but have not yet been fully characterised in terms of their composition, ratio of natural to synthetic component and membrane protein content. Here, we describe a method of correlative atomic force (AFM) with structured illumination microscopy (SIM) for the accurate mapping of complex lipid bilayers that consist of a synthetic fraction and a fraction of lipids derived from Escherichia coli outer membrane vesicles (OMVs). We exploit the enhanced resolution and molecular specificity that SIM can offer to identify areas of interest in these bilayers and the atomic scale resolution that the AFM provides to create detailed topography maps of the bilayers. We are thus able to understand the way in which the two different lipid fractions (natural and synthetic) mix within the bilayers, quantify the amount of bacterial membrane incorporated in the bilayer and directly visualise the interaction of these bilayers with bacteria-specific, membrane-binding proteins. Our work sets the foundation for accurately understanding the composition and properties of OMV-derived SLBs and establishes correlative AFM/ SIM as a method for characterising complex systems at the nanoscale.

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The Structural Integrity of the Model Lipid Membrane during Induced Lipid Peroxidation: The Role of Flavonols in the Inhibition of Lipid Peroxidation

Antioxidants ◽

10.3390/antiox9050430 ◽

2020 ◽

Vol 9 (5) ◽

pp. 430 ◽

Cited By ~ 2

Author(s):

Anja Sadžak ◽

Janez Mravljak ◽

Nadica Maltar-Strmečki ◽

Zoran Arsov ◽

Goran Baranović ◽

...

Keyword(s):

Lipid Peroxidation ◽

Lipid Bilayers ◽

Lipid Membranes ◽

Structural Changes ◽

Structural Integrity ◽

Lipid Membrane ◽

Membrane Properties ◽

Structural Reorganization ◽

Unsaturated Lipids ◽

Oxidative Attack

The structural integrity, elasticity, and fluidity of lipid membranes are critical for cellular activities such as communication between cells, exocytosis, and endocytosis. Unsaturated lipids, the main components of biological membranes, are particularly susceptible to the oxidative attack of reactive oxygen species. The peroxidation of unsaturated lipids, in our case 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), induces the structural reorganization of the membrane. We have employed a multi-technique approach to analyze typical properties of lipid bilayers, i.e., roughness, thickness, elasticity, and fluidity. We compared the alteration of the membrane properties upon initiated lipid peroxidation and examined the ability of flavonols, namely quercetin (QUE), myricetin (MCE), and myricitrin (MCI) at different molar fractions, to inhibit this change. Using Mass Spectrometry (MS) and Fourier Transform Infrared Spectroscopy (FTIR), we identified various carbonyl products and examined the extent of the reaction. From Atomic Force Microscopy (AFM), Force Spectroscopy (FS), Small Angle X-Ray Scattering (SAXS), and Electron Paramagnetic Resonance (EPR) experiments, we concluded that the membranes with inserted flavonols exhibit resistance against the structural changes induced by the oxidative attack, which is a finding with multiple biological implications. Our approach reveals the interplay between the flavonol molecular structure and the crucial membrane properties under oxidative attack and provides insight into the pathophysiology of cellular oxidative injury.

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The role of surface charge in the interaction of nanoparticles with model pulmonary surfactants

Soft Matter ◽

10.1039/c8sm00925b ◽

2018 ◽

Vol 14 (28) ◽

pp. 5764-5774 ◽

Cited By ~ 11

Author(s):

F. Mousseau ◽

J.-F. Berret

Keyword(s):

Surface Charge ◽

Lipid Bilayers ◽

Pulmonary Surfactant ◽

Lipid Vesicles ◽

Electrostatic Charge ◽

Supported Lipid Bilayers ◽

Pulmonary Surfactants ◽

Inhaled Nanoparticles

Inhaled nanoparticles reaching the respiratory zone in the lungs enter first in contact with the pulmonary surfactant. It is shown here that nanoparticles and lipid vesicles formulated from different surfactant mimetics interact predominantlyviaelectrostatic charge mediated attraction and do not form supported lipid bilayers spontaneously.

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Nanoscale imaging of bacterial infections by sphingolipid expansion microscopy

Nature Communications ◽

10.1038/s41467-020-19897-1 ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Ralph Götz ◽

Tobias C. Kunz ◽

Julian Fink ◽

Franziska Solger ◽

Jan Schlegel ◽

...

Keyword(s):

Lipid Bilayers ◽

Bacterial Infections ◽

Super Resolution ◽

Structured Illumination ◽

Structured Illumination Microscopy ◽

Nanoscale Imaging ◽

Intracellular Organelles ◽

Resolution Imaging ◽

Simkania Negevensis ◽

20 Nm

AbstractExpansion microscopy (ExM) enables super-resolution imaging of proteins and nucleic acids on conventional microscopes. However, imaging of details of the organization of lipid bilayers by light microscopy remains challenging. We introduce an unnatural short-chain azide- and amino-modified sphingolipid ceramide, which upon incorporation into membranes can be labeled by click chemistry and linked into hydrogels, followed by 4× to 10× expansion. Confocal and structured illumination microscopy (SIM) enable imaging of sphingolipids and their interactions with proteins in the plasma membrane and membrane of intracellular organelles with a spatial resolution of 10–20 nm. As our functionalized sphingolipids accumulate efficiently in pathogens, we use sphingolipid ExM to investigate bacterial infections of human HeLa229 cells by Neisseria gonorrhoeae, Chlamydia trachomatis and Simkania negevensis with a resolution so far only provided by electron microscopy. In particular, sphingolipid ExM allows us to visualize the inner and outer membrane of intracellular bacteria and determine their distance to 27.6 ± 7.7 nm.

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Characterization of Lipid Membrane Properties for Tunable Electroporation

ASME 2012 Third International Conference on Micro/Nanoscale Heat and Mass Transfer ◽

10.1115/mnhmt2012-75321 ◽

2012 ◽

Author(s):

H. Jeremy Cho ◽

Shalabh C. Maroo ◽

Evelyn N. Wang

Keyword(s):

Lipid Bilayers ◽

Lipid Membrane ◽

Contact Angle Measurement ◽

Angle Measurement ◽

Membrane Properties ◽

Force Microscopy ◽

Packing Structure ◽

Atomic Force ◽

Dynamics Simulations

Lipid bilayers form nanopores on the application of an electric field. This process of electroporation can be utilized in different applications ranging from targeted drug delivery in cells to nano-gating membrane for engineering applications. However, the ease of electroporation is dependent on the surface energy of the lipid layers and thus directly related to the packing structure of the lipid molecules. 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayers were deposited on a mica substrate using the Langmuir-Blodgett (LB) technique at different packing densities and analyzed using atomic force microscopy (AFM). The wetting behavior of these monolayers was investigated by contact angle measurement and molecular dynamics simulations. It was found that an equilibrium packing density of liquid-condensed (LC) phase DPPC likely exists and that water molecules can penetrate the monolayer displacing the lipid molecules. The surface tension of the monolayer in air and water was obtained along with its breakthrough force.

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Nanoparticle-Lipid Interaction: Job Scattering Plots to Differentiate Vesicle Aggregation from Supported Lipid Bilayer Formation

Colloids and Interfaces ◽

10.3390/colloids2040050 ◽

2018 ◽

Vol 2 (4) ◽

pp. 50 ◽

Cited By ~ 3

Author(s):

Fanny Mousseau ◽

Evdokia Oikonomou ◽

Victor Baldim ◽

Stéphane Mornet ◽

Jean-François Berret

Keyword(s):

Lipid Bilayers ◽

Lipid Vesicles ◽

Living Cells ◽

Supported Lipid Bilayers ◽

Supported Lipid Bilayer ◽

Mixed Aggregates ◽

Method Of Continuous Variation ◽

The Impact ◽

Analytical Expressions ◽

Lipid Interaction

The impact of nanomaterials on lung fluids, or on the plasma membrane of living cells, has prompted researchers to examine the interactions between nanoparticles and lipid vesicles. Recent studies have shown that nanoparticle-lipid interaction leads to a broad range of structures including supported lipid bilayers (SLB), particles adsorbed at the surface or internalized inside vesicles, and mixed aggregates. Currently, there is a need to have simple protocols that can readily evaluate the structures made from particles and vesicles. Here we apply the method of continuous variation for measuring Job scattering plots and provide analytical expressions for the scattering intensity in various scenarios. The result that emerges from the comparison between experiments and modeling is that electrostatics play a key role in the association, but it is not sufficient to induce the formation of supported lipid bilayers.

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Site-Specific DNA-Controlled Fusion of Single Lipid Vesicles to Supported Lipid Bilayers

Biophysical Journal ◽

10.1016/j.bpj.2009.12.3698 ◽

2010 ◽

Vol 98 (3) ◽

pp. 673a

Author(s):

Lisa V. Simonsson ◽

Peter Jönsson ◽

Gudrun Stengel ◽

Fredrik Höök

Keyword(s):

Lipid Bilayers ◽

Lipid Vesicles ◽

Supported Lipid Bilayers ◽

Site Specific ◽

Controlled Fusion

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Broadband Plasmon Waveguide Resonance Spectroscopy for Probing Biological Thin Films

Applied Spectroscopy ◽

10.1366/000370209789379295 ◽

2009 ◽

Vol 63 (9) ◽

pp. 1062-1067 ◽

Cited By ~ 11

Author(s):

Han Zhang ◽

Kristina S. Orosz ◽

Hiromi Takahashi ◽

S. Scott Saavedra

Keyword(s):

Lipid Bilayers ◽

Lipid Membrane ◽

Resonance Wavelength ◽

Supported Lipid Bilayers ◽

Resonance Spectroscopy ◽

Silica Layer ◽

Transmembrane Receptors ◽

Bk7 Glass ◽

Biological Thin Films ◽

Waveguide Resonance

A commercially available spectrometer has been modified to perform plasmon waveguide resonance (PWR) spectroscopy over a broad spectral bandwidth. When compared to surface plasmon resonance (SPR), PWR has the advantage of allowing measurements in both s- and p-polarizations on a waveguide surface that is silica or glass rather than a noble metal. Here the waveguide is a BK7 glass slide coated with silver and silica layers. The resonance wavelength is sensitive to the optical thickness of the medium adjacent to the silica layer. The sensitivity of this technique is characterized and compared with broadband SPR both experimentally and theoretically. The sensitivity of spectral PWR is comparable to that of spectral SPR for samples with refractive indices close to that of water. The hydrophilic surface of the waveguide allows supported lipid bilayers to be formed spontaneously by vesicle fusion; in contrast, the surface of an SPR chip requires chemical modification to create a supported lipid membrane. Broadband PWR spectroscopy should be a useful technique to study biointerfaces, including ligand binding to transmembrane receptors and adsorption of peripheral proteins on ligand-bearing membranes.

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Adhesion and Structural Changes of PEGylated Lipid Nanocarriers on Silica Surfaces

Physchem ◽

10.3390/physchem1020009 ◽

2021 ◽

Vol 1 (2) ◽

pp. 133-151

Author(s):

Philipp Grad ◽

Katarina Edwards ◽

Víctor Agmo Hernández

Keyword(s):

Lipid Bilayers ◽

Structural Changes ◽

Lipid Membrane ◽

Complete Separation ◽

Supported Lipid Bilayers ◽

Pegylated Liposomes ◽

Silica Surfaces ◽

Lipid Nanocarriers ◽

Liquid Ordered ◽

Pegylated Lipids

PEGylated lipid nanoparticles have a continuously expanding range of applications, particularly within pharmaceutical areas. Hereby, it is shown with the help of the Quartz Crystal Microbalance with Dissipation monitoring (QCM-D) and other surface sensitive techniques that, at room temperature, PEGylated liposomes and lipodisks adhere strongly to silica surfaces resulting in the displacement of the hydration layer of silica and the formation of immobilized nanoparticle films. Furthermore, it is shown that drastic changes in the structure of the immobilized films occur if the temperature is increased to >35 °C. Thus, intact immobilized PEGylated liposomes rupture and spread, even in the gel phase state; immobilized lipodisks undergo complete separation of their components (bilayer forming lipids and PEGylated lipids) resulting in a monolayer of adsorbed PEGylated lipids; and PEGylated supported lipid bilayers release part of the water trapped between the lipid membrane and the surface. It is hypothesized that these changes occur mainly due to the changes in the configuration of PEG chains and a drastic decrease of the affinity of the polymer for water. The observed phenomena can be applied, e.g., for the production of defect-free supported lipid bilayers in the gel or liquid ordered phase states.

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Nanoparticle-Lipid Interaction: Job Scattering Plots to Differentiate Vesicle Aggregationfrom Supported Lipid Bilayer Formation

10.20944/preprints201809.0421.v1 ◽

2018 ◽

Author(s):

Fanny Mousseau ◽

Evdokia Oikonomou ◽

Victor Baldim ◽

Stéphane Mornet ◽

Jean-François Berret

Keyword(s):

Lipid Bilayers ◽

Lipid Vesicles ◽

Living Cells ◽

Supported Lipid Bilayers ◽

Supported Lipid Bilayer ◽

Mixed Aggregates ◽

Method Of Continuous Variation ◽

The Impact ◽

Analytical Expressions ◽

Lipid Interaction

The impact of nanomaterials on lung fluids or on the plasma membrane of living cells has prompted researchers to examine the interactions between nanoparticles and lipid vesicles. Recent studies have shown that nanoparticle-lipid interaction leads to a broad range of structures including supported lipid bilayers (SLB), particles adsorbed at the surface or internalized inside vesicles, and mixed aggregates. Today, there is a need to have simple protocols that can readily assess the nature of structures obtained from particles and vesicles. Here we apply the method of continuous variation for measuring Job scattering plots and provide analytical expressions for the scattering intensity in various scenarios. The result that emerges from the comparison between modeling and experimental measurements is that electrostatics plays a key role in the association, but it is not sufficient to induce the formation of supported lipid bilayers.

Download Full-text