Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 9 de 9
Filter
Add more filters










Database
Language
Publication year range
1.
PLoS Pathog ; 14(1): e1006814, 2018 01.
Article in English | MEDLINE | ID: mdl-29320578

ABSTRACT

Mycolactone is a lipid-like endotoxin synthesized by an environmental human pathogen, Mycobacterium ulcerans, the causal agent of Buruli ulcer disease. Mycolactone has pleiotropic effects on fundamental cellular processes (cell adhesion, cell death and inflammation). Various cellular targets of mycolactone have been identified and a literature survey revealed that most of these targets are membrane receptors residing in ordered plasma membrane nanodomains, within which their functionalities can be modulated. We investigated the capacity of mycolactone to interact with membranes, to evaluate its effects on membrane lipid organization following its diffusion across the cell membrane. We used Langmuir monolayers as a cell membrane model. Experiments were carried out with a lipid composition chosen to be as similar as possible to that of the plasma membrane. Mycolactone, which has surfactant properties, with an apparent saturation concentration of 1 µM, interacted with the membrane at very low concentrations (60 nM). The interaction of mycolactone with the membrane was mediated by the presence of cholesterol and, like detergents, mycolactone reshaped the membrane. In its monomeric form, this toxin modifies lipid segregation in the monolayer, strongly affecting the formation of ordered microdomains. These findings suggest that mycolactone disturbs lipid organization in the biological membranes it crosses, with potential effects on cell functions and signaling pathways. Microdomain remodeling may therefore underlie molecular events, accounting for the ability of mycolactone to attack multiple targets and providing new insight into a single unifying mechanism underlying the pleiotropic effects of this molecule. This membrane remodeling may act in synergy with the other known effects of mycolactone on its intracellular targets, potentiating these effects.


Subject(s)
Lipid Bilayers , Macrolides/pharmacology , Membrane Microdomains/drug effects , Buruli Ulcer/microbiology , Cell Adhesion/drug effects , Humans , Lipid Bilayers/chemistry , Lipid Bilayers/metabolism , Membrane Lipids/chemistry , Membrane Lipids/metabolism , Membrane Microdomains/metabolism , Microbial Sensitivity Tests , Mycobacterium ulcerans/chemistry , Mycobacterium ulcerans/drug effects , Mycobacterium ulcerans/ultrastructure , Surface-Active Agents/pharmacology
2.
Biochim Biophys Acta ; 1838(10): 2698-707, 2014 Oct.
Article in English | MEDLINE | ID: mdl-25019684

ABSTRACT

VIsinin-LIke Proteins (VILIPs) are a subfamily of the Neuronal Calcium Sensor (NCS) proteins, which possess both N-myristoylation and EF-hand motifs allowing for a putative 'calcium-myristoyl switch' regulation mechanism. It has previously been established that myristoyl conjugation increases the affinity of proteins for membranes, but, in many cases, a second feature such as a cluster of positively-charged residues is needed for stable membrane binding. The interaction of two members of this family, VILIP-1 and VILIP-3, with Langmuir monolayers as membrane models has been investigated in order to study the effects of both myristoylation and the highly basic region containing conserved poly-lysine residues on membrane association kinetics and binding properties. Results show that in the presence of calcium, N-myristoylation significantly increases the kinetic rate of VILIP adsorption to the membrane. Additionally, the proteins bind to negatively charged phospholipids independently of the conjugated myristate moiety. Besides the regulatory effect of calcium on the rate of binding presumably due to exposure of the myristoyl moiety ascribed to their putative 'calcium-myristoyl switch', VILIP-1 and -3 also engage specific interactions with biomimetic membranes containing phosphatidylinositol 4,5-bisphosphate (PIP2). The presence of PIP2 increases the membrane association rates of both VILIPs. Taken together, these results show the major kinetic role of N-myristoylation for membrane binding, and highlight the critical role of specific phosphoinositide interactions for membrane association of members of the VILIP family.


Subject(s)
Calcium/metabolism , Cell Membrane/metabolism , Lipoylation , Membranes, Artificial , Neurocalcin/metabolism , Phosphatidylinositol Phosphates/metabolism , Calcium/chemistry , Cell Membrane/chemistry , Humans , Neurocalcin/chemistry , Phosphatidylinositol Phosphates/chemistry
3.
Biochimie ; 107 Pt A: 135-42, 2014 Dec.
Article in English | MEDLINE | ID: mdl-24998327

ABSTRACT

Biological membranes play a central role in the biology of the cell. They are not only the hydrophobic barrier allowing separation between two water soluble compartments but also a supra-molecular entity that has vital structural functions. Notably, they are involved in many exchange processes between the outside and inside cellular spaces. Accounting for the complexity of cell membranes, reliable models are needed to acquire current knowledge of the molecular processes occurring in membranes. To simplify the investigation of lipid/protein interactions, the use of biomimetic membranes is an approach that allows manipulation of the lipid composition of specific domains and/or the protein composition, and the evaluation of the reciprocal effects. Since the middle of the 80's, lipid bilayer membranes have been constantly developed as models of biological membranes with the ultimate goal to reincorporate membrane proteins for their functional investigation. In this review, after a brief description of the planar lipid bilayers as biomimetic membrane models, we will focus on the construction of the tethered Bilayer Lipid Membranes, the most promising model for efficient membrane protein reconstitution and investigation of molecular processes occurring in cell membranes.


Subject(s)
Cell Membrane/metabolism , Lipid Bilayers/metabolism , Membrane Lipids/metabolism , Membrane Proteins/metabolism , Amino Acid Sequence , Cell Membrane/chemistry , Lipid Bilayers/chemistry , Liposomes/chemistry , Liposomes/metabolism , Membrane Lipids/chemistry , Membrane Proteins/chemistry , Models, Molecular , Molecular Sequence Data , Peptides/chemistry , Peptides/metabolism , Protein Binding
4.
Anal Chem ; 82(6): 2401-4, 2010 Mar 15.
Article in English | MEDLINE | ID: mdl-20163148

ABSTRACT

The effect of the ionic liquid 1-ethyl-3-methylimidazolium ethylsulfate ([Emim][EtSO(4)]) on the copper-catalyzed luminol chemiluminescence (CL) is reported. A drastic light emission enhancement is observed, related to a strong interaction between Cu(2+) and the imidazolium ring. In these conditions, the CL reaction was able to produce light efficiently at pH as low as 6.5 (amplification factor: Intensity(+IL)/Intensity(-IL) = 2900). Interesting effects of [Emim][EtSO(4)] on the enzyme glucose oxidase activity were also evidenced, and advantages were taken from this enhancement to perform sensitive chemiluminescent glucose detection (LOD = 4 microM) at pH 8.0.


Subject(s)
Copper/chemistry , Glucose/analysis , Imidazoles/chemistry , Luminescent Measurements/methods , Aspergillus niger/enzymology , Catalysis , Glucose Oxidase/metabolism , Hydrogen Peroxide/analysis , Limit of Detection
5.
Langmuir ; 26(3): 2160-6, 2010 Feb 02.
Article in English | MEDLINE | ID: mdl-20000740

ABSTRACT

The present study reports the achievement of a new chemiluminescent sensing layer able to simultaneously (i) play an active role on ligand immobilization and (ii) serve as a catalyst in detection processes for label-free biosensor applications. This new type of active Langmuir-Blodgett (LB) monolayer has been designed by using a chelating lipid (Ni-NTA-DOGS). Thanks to the chelated metallic cation, this peculiar lipid exhibits luminol chemiluminescence catalysis properties in the presence of hydrogen peroxide. Upon biomolecule interaction through imidazole ring chelation (mediated by the metallic cation bound to the lipid headgroup), the chemiluminescent signal can be modulated. The first chemiluminescent signal acquisition experiments have shown a strong and homogeneous signal of the chelating layer. Upon histamine interactions, a histidine derivative used as a marker of fresh food quality, we succeeded in obtaining as a proof of concept a chemiluminescent signal variation without any derivatization of the target molecule. This signal variation was shown to be directly correlated to the histamine concentration with a limit of detection of 2 microg/mL.


Subject(s)
Biosensing Techniques/methods , Chelating Agents/chemistry , Catalysis , Copper/chemistry , Histamine/analysis , Histamine/chemistry , Histamine/metabolism , Immobilized Proteins/chemistry , Immobilized Proteins/metabolism , Lipid Metabolism , Luminescent Measurements , Luminol/chemistry , Lysine/analogs & derivatives , Lysine/chemistry , Nanotechnology , Nickel/chemistry , Oleic Acids/chemistry , Succinates/chemistry , Surface Properties , Water/chemistry
6.
Adv Colloid Interface Sci ; 116(1-3): 205-25, 2005 Nov 30.
Article in English | MEDLINE | ID: mdl-16181605

ABSTRACT

This review presents the recent advances in the achievement of organized proteo-lipidic nanostructures based on Langmuir-Blodgett technology and their potential applications in the nanobioscience area. By using the self-assembled properties of amphiphilic biomolecules at the air-water interface, the Langmuir-Blodgett (LB) technique offers the possibility to prepare ultrathin layers suitable for biomolecule immobilization at the molecular level. This review will provide a general overview of the enzyme association with preformed Langmuir-Blodgett films in connection with their potential applications in biosensing device developments, and then introduce the design of a new functionalised biomimetic nanostructure with oriented recognition site. The potential applications of such an organized proteo-lipidic nanostructure for biocatalysis investigations of an immobilised enzyme in a biomimetic situation and for the development of bioelectronic devices are finally discussed.


Subject(s)
Enzymes/chemistry , Lipids/chemistry , Membranes, Artificial , Nanotechnology/instrumentation , Nanotechnology/methods , Models, Biological , Surface Properties
7.
Biosens Bioelectron ; 20(8): 1539-48, 2005 Feb 15.
Article in English | MEDLINE | ID: mdl-15626607

ABSTRACT

This work presents a new approach for direct detection of polyelectrolytes at the air-water interface, based on the investigation of the interfacial properties of an active lipidic matrix especially designed for polynucleotide immobilization. A synthetic lipid with a cationic spermine headgroup, DiOctadecylamidoGlycylSpermine (DOGS), was spread at the interface to form a distortable film able to capture polynucleotides. The control of the organization state of this functionalized monolayer upon compression was achieved by recording surface pressure-area (pi-A) isotherm diagrams, presenting a specific shape with a typical liquid expanded-liquid condensed phase transition on a pure water subphase. In the presence of various dsDNA concentrations in the subphase, the isotherms were markedly modified in a time and concentration-dependent manner. The main modifications, corresponding to a large shift towards higher molecular areas and a clear fading of the phase transition, were corroborated by the fine analysis of the monolayer compressibility profile, thus suggesting a characteristic change in the monolayer fluidity as a function of both time and DNA concentration. Moreover, an ATR-Fourier transform infrared (ATR-FTIR) characterization showed evidences for the adsorption of DNA strands onto the lipidic matrix. The direct observation of the mixed monolayer morphology by Brewster angle microscopy (BAM) strongly suggests that DNA adsorption induces a reorganization of lipids at the interface, as evidenced by the change in the condensed lipidic domains morphology in the presence of DNA in the subphase. The immobilization of various polynucleotidic probes of 4000, 400 and 22 base length, confirmed by fluorescence microscopy, had similar effects on DOGS interfacial properties. Preliminary studies are finally presented to explore the possibility of using this system for the study of hybridization between complementary strands. Hence, this study demonstrates this functionalized matrix behaves as a fluid support where polynucleotide immobilization induces interfacial properties modifications, which could be further exploited through the experimental characterization of Faraday instabilities sensitive to visco-elasticity variations.


Subject(s)
Biosensing Techniques/methods , DNA/analysis , In Situ Hybridization/methods , Microfluidic Analytical Techniques/methods , Oligonucleotide Probes/analysis , Polynucleotides/analysis , Spermidine/antagonists & inhibitors , Spermidine/analysis , Adsorption , Biosensing Techniques/instrumentation , DNA/chemistry , DNA/genetics , In Situ Hybridization/instrumentation , Membrane Lipids/analysis , Membrane Lipids/chemistry , Membranes, Artificial , Microfluidic Analytical Techniques/instrumentation , Oligonucleotide Array Sequence Analysis/instrumentation , Oligonucleotide Array Sequence Analysis/methods , Oligonucleotide Probes/chemistry , Polynucleotides/chemistry , Spermidine/chemistry , Surface Tension
8.
Chembiochem ; 6(2): 395-404, 2005 Feb.
Article in English | MEDLINE | ID: mdl-15651043

ABSTRACT

This study deals with the kinetics properties of an enzyme immobilised in a defined orientation in a biomimetic environment. For this purpose, acetylcholinesterase (AChE) was captured at the surface of a nanostructured proteo-glycolipidic Langmuir-Blodgett film through specific recognition by a noninhibitor monoclonal antibody (IgG) inserted in a neoglycolipid bilayer. Modelling of this molecular assembly provided a plausible interpretation of the functional orientation of the enzyme. The AChE activity being stable for several weeks, the enzyme kinetics were investigated, and fitted perfectly with heterogeneous biocatalytic behaviour representative of cellular enzymatic catalysis. The AChE-IgG-glycolipid nanostructure was directly interfaced with an efficient optical device. Such an association, leading to an intimate contact between the nanostructure and the biochemical signal transducer, gives direct access to the intrinsic AChE behaviour. This study thus demonstrates the potential for direct investigation of the kinetic behaviour of an immobilised enzyme on a lipid bilayer through an efficient transduction system.


Subject(s)
Acetylcholinesterase/pharmacokinetics , Bungarus , Enzymes, Immobilized/pharmacokinetics , Lipid Bilayers , Snake Venoms/enzymology , Acetylcholinesterase/chemistry , Amino Acid Sequence , Animals , Enzyme Stability , Enzymes, Immobilized/chemistry , Luminescent Measurements , Models, Molecular , Molecular Sequence Data , Molecular Structure , Protein Conformation , Proteolipids/chemistry , Proteolipids/metabolism , Sequence Alignment , Snake Venoms/chemistry
9.
Biochim Biophys Acta ; 1617(1-2): 39-51, 2003 Oct 31.
Article in English | MEDLINE | ID: mdl-14637018

ABSTRACT

The insertion of immunoglobulin (IgG) in a glycolipid monolayer was achieved by using the ability of new proteo-glycolipid vesicles to disintegrate into a mixed IgG-glycolipid interfacial film after spreading at an air-buffer interface. The interfacial disintegration kinetics was shown to be directly dependent on the initial vesicle surface density and on the buffer ionic strength. The presence of the immunoglobulin in the glycolipid film was displayed by an increase of the lateral compressibility (Cs) during monolayer compression. Cs magnitude modifications, due to the antibody effect on the monolayer packing, decreases as the spread vesicle density increases. At interfacial saturation, the lateral compressibility profile becomes similar to that of a control monolayer without antibody. However, the careful analysis of the mixed monolayer after transfer by Langmuir-Blodgett technique (ATR-FTIR characterisation, enzyme immunoassociation) clearly demonstrated that the antibody was still present in such conditions and was not completely squeezed out from the interface as compressibility changes could have meant. At nonsaturating vesicle surface density, IgG molecules initially lying in the lipid matrix with the Y-shape plane parallel to the interface move to a standing-up position during the compression, leading to lateral compressibility modifications. For a saturating vesicle surface density, the glycolipid molecules force the IgG molecules to directly adopt a more vertical position in the interfacial film and, consequently, no lateral compressibility modification was recorded during the compression.


Subject(s)
Biocompatible Materials/chemical synthesis , Glycolipids/chemistry , Immunoglobulin G/chemistry , Liposomes/chemistry , Membrane Fluidity , Air , Biocompatible Materials/chemistry , Biosensing Techniques , Buffers , Diffusion , Immunoglobulin G/immunology , Kinetics , Liposomes/chemical synthesis , Membranes, Artificial , Protein Conformation , Solutions , Surface Properties , Surface Tension
SELECTION OF CITATIONS
SEARCH DETAIL
...