Publications

Année Type Filtrer >


1 […]
GAITAN, Julien; MILOCHAU, Alexandra; LANG, Jochen;
Loss-of-function mutations in ADCY3 cause monogenic severe obesity
Nat Genet, 2018, 0,
Study of monogenic forms of obesity has demonstrated the pivotal role of the central leptin-melanocortin pathway in controlling energy balance, appetite and body weight (1) . The majority of loss-of-function mutations (mostly recessive or co-dominant) have been identified in genes that are directly involved in leptin-melanocortin signaling. These genes, however, only explain obesity in <5% of cases, predominantly from outbred populations (2) . We previously showed that, in a consanguineous population in Pakistan, recessive mutations in known obesity-related genes explain ~30% of cases with severe obesity(3-5). These data suggested that new monogenic forms of obesity could also be identified in this population. Here we identify and functionally characterize homozygous mutations in the ADCY3 gene encoding adenylate cyclase 3 in children with severe obesity from consanguineous Pakistani families, as well as compound heterozygous mutations in a severely obese child of European-American descent. These findings highlight ADCY3 as an important mediator of energy homeostasis and an attractive pharmacological target in the treatment of obesity.
2 […]
ZHANG, Jiarong; BATHANY, Katell; CHAUDIERE, Jean;
Production of 3,4-cis- and 3,4-trans-Leucocyanidin and Their Distinct MS/MS Fragmentation Patterns
Journal of Agricultural and Food Chemistry, 2018, 66, 351-358
(+)-2,3-trans-3,4-cis-Leucocyanidin was produced by acidic epimerization of (+)-2,3-trans-3,4-trans-leucocyanidin synthesized by reduction of (+)-dihydroquercetin with NaBH4, and structures of the two stereoisomers purified by C18- and phenyl-reverse-phase high-performance liquid chromatography (HPLC) were confirmed by NMR spectroscopy. We confirm that only 3,4-cis-leucocyanidin is used by leucoanthocyanidin reductase as substrate. The two stereoisomers are quite stable in aqueous solution at -20 degrees C. Characterization of the two stereoisomers was also performed using electrospray ionization tandem mass spectrometry (ESI-MS/MS), and we discuss here for the first time the corresponding MS/MS fragmentation pathways, which are clearly distinct. The main difference is that of the mode of dehydration of the 3,4-diol in positive ionization mode, which involves a loss of hydroxyl group at either C-3 or C-4 for the 3,4-cis isomer but only at C-3 for the 3,4-trans isomer. Tandem mass spectrometry therefore proves useful as a complementary methodology to NMR to identify each of the two stereoisomers.
3 […]
GENESTE, Emmanuel; GRELARD, Axelle; DUFOURC, Erick;
Photodegradation of novel oral anticoagulants under sunlight irradiation in aqueous matrices
Chemosphere, 2018, 193, 329-336
Kinetics of photodegradation of novel oral anticoagulants dabigatran, rivaroxaban, and apixaban were studied under simulated solar light irradiation in purified, mineral, and river waters. Dabigatran and rivaroxaban underwent direct photolysis with polychromatic quantum yields of 2.2 x 10(-4) and 4.4 x 10(-2), respectively. The direct photodegradation of apixaban was not observed after 19 h of irradiation. Kinetics of degradation of rivaroxaban was not impacted by the nature of the aqueous matrix while photosensitization from nitrate ions was observed for dabigatran and apixaban dissolved in a mineral water. The photosensitized reactions were limited in the tested river water (Isle River, Perigueux, France) certainly due to the hydroxyl radical scavenging effect of the dissolved organic matter. The study of photoproduct structures allowed to identify two compounds for dabigatran. One of them is the 4-aminobenzamidine while the second one is a cyclization product. In the case of rivaroxaban, as studied by very high field NMR, only one photoproduct was observed i.e. a photoisomer. Finally, seven photoproducts were clearly identified from the degradation of apixaban under simulated solar light. (C) 2017 Elsevier Ltd. All rights reserved.
4 […]
SEHL, Anthony; COUEDELO, Leslie; CANSELL, Maud;
A critical assessment of transmethylation procedures for n-3 long-chain polyunsaturated fatty acid quantification of lipid classes
Food chemistry, 2018, 251, 1-8
Lipid transmethylation methods described in the literature are not always evaluated with care so to insure that the methods are effective, especially on food matrix or biological samples containing polyunsaturated fatty acid (PUFA). The aim of the present study was to select a method suitable for all lipid species rich in long chain n-3 PUFA. Three published methods were adapted and applied on individual lipid classes. Lipid (trans)methylation efficiency was characterized in terms of reaction yield and gas chromatography (GC) analysis. The acid-catalyzed method was unable to convert triglycerides and sterol esters, while the method using an incubation at a moderate temperature was ineffective on phospholipids and sterol esters. On the whole only the method using sodium methoxide and sulfuric acid was effective on lipid classes taken individually or in a complex medium. This study highlighted the use of an appropriate (trans)methylation method for insuring an accurate fatty acid composition. Copyright © 2018 Elsevier Ltd. All rights reserved.
5 […]
SAAD, Ahmad; LIET, Benjamin; JOUCLA, Gilles; SANTARELLI, Xavier; TREZEGUET, Véronique;
Role of Glycanation and Convertase Maturation of Soluble Glypican-3 in Inhibiting Proliferation of Hepatocellular Carcinoma Cells
Biochemistry, 2018, 0,
Glypican 3 (GPC3) is a complex heparan sulfate proteoglycan associated with the outer surface of the plasma membrane by a glycosylphosphatidylinositol (GPI) anchor. It is also N-glycosylated and processed by a furin-like convertase. GPC3 has numerous biological functions. Although GPC3 is undetectable in normal liver tissue, it is abnormally and highly overexpressed in hepatocellular carcinoma (HCC). Interestingly, proliferation of HCC cells such as HepG2 and HuH7 is inhibited when they express a soluble form of GPC3 after lentiviral transduction. To obtain more insight into the role of some of its post-translational modifications, we designed a mutant GPC3, sGPC3m, without its GPI anchor, convertase cleavage site, and glycosaminoglycan chains. The highly pure sGPC3m protein strongly inhibited HuH7 and HepG2 cell proliferation in vitro and induced a significant increase in their cell doubling time. It changed the morphology of HuH7 cells but not that of HepG2. It induced the enlargement of HuH7 cell nuclear area and the restructuration of adherent cell junctions. Unexpectedly, for both cell types, the levels of apoptosis, cell division, and beta-catenin were not altered by sGPC3m, although growth inhibition was very efficient. Overall, our data show that glycanation and convertase maturation are not required for sGPC3m to inhibit HCC cell proliferation.
6 […]
ANTUNES, Stéphanie; PERDRIAU, Camille; KAUFFMANN, Brice; PASCO, Morgane; DOUAT, Céline; GUICHARD, Gilles;
Postelongation Strategy for the Introduction of Guanidinium Units in the Main Chain of Helical Oligourea Foldamers
The Journal of organic chemistry, 2018, 0,
The synthesis of hybrid urea-based foldamers containing isosteric guanidinium linkages at selected positions in the sequence is described. We used a postelongation approach whereby the guanidinium moiety is introduced by direct transformation of a parent oligo(urea/thiourea) foldamer precursor. The method involves activation of the thiourea by treatment with methyl iodide and subsequent reaction with amines. To avoid undesired cyclization with the preceding urea moiety, resulting in heterocyclic guanidinium formation in the main chain, the urea unit preceding the thiourea unit in the sequence was replaced by an isoatomic and isostructural gamma-amino acid. The approach was extended to solid-phase techniques to accelerate the synthesis of longer and more functionalized sequences. Under optimized conditions, an octamer hybrid oligomer incorporating a central guanidinium linkage was obtained in good overall yield and purity. This work also reports data related to the structural consequences of urea by guanidinium replacements in solution and reveals that helical folding is substantially reduced in oligomers containing a guanidinium group.
7 […]
GEAN, Julie; DUFOURC, Erick;
The potent effect of mycolactone on lipid membranes
Plos Pathogens, 2018, 14,
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 mu M, 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.
8 […]
SCHMITTER, Jean-Marie;
Comprehensive targeted and non-targeted lipidomics analyses in failing and non-failing heart
Analytical and bioanalytical chemistry, 2018, 0,
Myocardial infarction (MI) and subsequent progressive heart failure pathology is the major cause of death worldwide; however, the mechanism of this pathology remains unclear. The present work aimed at testing the hypothesis whether the inflammatory response is superimposed with the formation of bioactive lipid resolving molecules at the site of the injured myocardium in acute heart failure pathology post-MI. In this view, we used a robust permanent coronary ligation model to induce MI, leading to decreased contractility index with marked wall thinning and necrosis of the infarcted left ventricle. Then, we applied mass spectrometry imaging (MSI) in positive and negative ionization modes to characterize the spatial distribution of left ventricle lipids in the infarcted myocardium post-MI. After micro-extraction, liquid chromatography coupled to tandem mass spectrometry was used to confirm the structures of the imaged lipids. Statistical tools such as principal component analysis were used to establish a comprehensive visualization of lipid profile changes in MI and no-MI hearts. Resolving bioactive molecules such as resolvin (Rv) D1, RvD5, RvE3, 17-HDHA, LXA4, and 18-HEPE were detected in negative ion mode MSI, whereas phosphatidyl cholines (PC) and oxidized derivatives thereof were detected in positive ion mode. MSI-based analysis demonstrated a significant increase in resolvin bioactive lipids with comprehensive lipid remodeling at the site of infarction. These results clearly indicate that infarcted myocardium is the primary location of inflammation-resolution pathomechanics which is critical for resolution of inflammation and heart failure pathophysiology. Graphical abstract Applied scheme to determine comprehensive lipidomics in failing and non-failing heart.
9 […]
HARTE, Etienne; ELEZGARAY, Juan; ALVES, Isabel; LECOMTE, Sophie;
DISPOSITIF ET PROCEDE POUR LA CARACTERISATION D'ECHANTILLONS PAR IMAGERIE DE SPECTROSCOPIE PWR
, 2018, 0,
La présente invention concerne un dispositif et un procédé pour la mesure par spectroscopie d'un échantillon dans lequel on réalise les étapes suivantes: - envoyer un faisceau lumineux vers une surface (12) d'incidence d'un capteur optique, ledit faisceau lumineux formant un angle d'incidence avec ladite surface d'incidence, ledit faisceau lumineux comportant des ondes électromagnétiques p-polarisées et des ondes électromagnétiques s-polarisées, - ledit capteur optique comprenant un prisme (13) comportant une surface de réflexion, un premier film (15) conducteur ou semi-conducteur et un deuxième film (16) diélectrique pour générer deux modes guidés, un premier desdits modes guidés étant généré pour des ondes électromagnétiques p-polarisées et pour un premier angle d'incidence Qi dudit faisceau lumineux sur ladite surface (12) d'incidence et un second desdits modes guidés étant généré pour les ondes électromagnétiques s-polarisés et pour un second angle d'incidence
10 […]
DE, Soumen; CHI, Bo; GRANIER, Thierry; QI, Ting; MAURIZOT, Victor; HUC, Ivan;
Designing cooperatively folded abiotic uni- and multimolecular helix bundles
Nature Chemistry, 2018, 10, 51-57
Abiotic foldamers, that is foldamers that have backbones chemically remote from peptidic and nucleotidic skeletons, may give access to shapes and functions different to those of peptides and nucleotides. However, design methodologies towards abiotic tertiary and quaternary structures are yet to be developed. Here we report rationally designed interactional patterns to guide the folding and assembly of abiotic helix bundles. Computational design facilitated the introduction of hydrogen-bonding functionalities at defined locations on the aromatic amide backbones that promote cooperative folding into helix-turn-helix motifs in organic solvents. The hydrogen-bond-directed aggregation of helices not linked by a turn unit produced several thermodynamically and kinetically stable homochiral dimeric and trimeric bundles with structures that are distinct from the designed helix-turn-helix. Relative helix orientation within the bundles may be changed from parallel to tilted on subtle solvent variations. Altogether, these results prefigure the richness and uniqueness of abiotic tertiary structure behaviour.
11 […]
LECOMTE, Sophie;
Interfacial activity and emulsifying behaviour of inclusion complexes between helical polysaccharides and flavouring molecules resulting from non-covalent interactions
Food research international (Ottawa, Ont.), 2018, 105, 801-811
This study deals with the fabrication of inclusion complexes starting from a cross coupling of seven helical polysaccharides (host) and six flavouring agents (guest). Neither of the substrates is considered as an emulsifier when studied alone. Due to a complexation mechanism, the presence of intermolecular hydrogen bonds between substrates was highlighted by infra-red spectroscopy and 13C NMR. In addition, depending on the polysaccharide used, the guest molecule could be preferentially located either inside or in the interstitial spaces of the helix. In a comparison between raw substrates, the inclusion complexes obtained presented the unique interfacial activity of decreasing surface tension values (gamma) and, in some cases, their behaviour in water was similar to that of regular emulsifiers due to the presence of a critical aggregation concentration (CAC). Substrate concentrations and the ratios between them were the main parameters investigated in this study, which focused on the two inclusion complexes: vanillin/amylose and vanillin/iota-carrageenan. The first decreased gamma values by as much as 53mN/m with a double transition, whereas the second could cause gamma fall to 36mN/m with a regular break. In addition, these systems were able to stabilize foams for up to 60min, which confirmed their unique emulsifying properties.
12 […]
DALLET, Laurence; TAVEAU, Jean-Christophe; LECOMTE, Sophie; POUSSARD, Sylvie; LAMBERT, Olivier;
Single lipoaminoglycoside promotes efficient intracellular antibody delivery: A comprehensive insight into the mechanism of action
Nanomedicine-Nanotechnology Biology and Medicine, 2018, 14, 141-151
Delivery of biologically active proteins into cells is emerging as important strategy for many applications. Previous experiments have shown that lipoaminoglycosides were capable of delivery of the anti-cytokeratin8 antibody (anti-K8) but only when formulated with lipid helpers potentially leading to toxicity from excess lipids. Here, we optimized anti-K8 delivery with various lipoaminoglycosides in the absence of a lipid helper. Results led to the identification of the aminoglycoside lipid dioleyl phosphoramido ribostamycin (DOPRI) as a potent intracellular delivery system for anti-K8. Electron microscopy revealed that delivered anti-K8 molecules were bound to intermediate filaments in cells. Anti-K8 was bound to the surface of DOPRI vesicles without perturbing lipid organization. Macropinocytosis and caveolin mediated endocytosis contributed to anti-K8 internalization and to filament labeling with a major contribution being made by the caveolin pathway. The results showed that the unique properties of DOPRI were sufficient for efficient intracellular protein delivery without requiring lipid helpers. (C) 2017 Elsevier Inc. All rights reserved.
13 […]
CULLIN, Christophe;
Guidelines and recommendations on yeast cell death nomenclature
Microbial Cell, 2018, 5, 4-31
Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cellular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the definition of accidental, regulated, and programmed forms of cell death in yeast based on a series of morphological and biochemical criteria. Specifically, we provide consensus guidelines on the differential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death routines that are relevant for the biology of (at least some species of) yeast. As this area of investigation advances rapidly, changes and extensions to this set of recommendations will be implemented in the years to come. Nonetheless, we strongly encourage the authors, reviewers and editors of scientific articles to adopt these collective standards in order to establish an accurate framework for yeast cell death research and, ultimately, to accelerate the progress of this vibrant field of research.
14 […]
PLAWINSKI, Laurent; DURRIEU, Marie-Christine;
Opto-acoustic microscopy reveals adhesion mechanics of single cells
The Review of scientific instruments, 2018, 89, 014901-014901
Laser-generated GHz-ultrasonic-based technologies have shown the ability to image single cell adhesion and stiffness simultaneously. Using this new modality, we here demonstrate quantitative indicators to investigate contact mechanics and adhesion processes of the cell. We cultured human cells on a rigid substrate, and we used an inverted pulsed opto-acoustic microscope to generate acoustic pulses containing frequencies up to 100 GHz in the substrate. We map the reflection of the acoustic pulses at the cell-substrate interface to obtain images of the acoustic impedance of the cell, Zc, as well as of the stiffness of the interface, K, with 1 mum lateral resolution. Our results show that the standard deviation DeltaZc reveals differences between different cell types arising from the multiplicity of local conformations within the nucleus. From the distribution of K-values within the nuclear region, we extract a mean interfacial stiffness, Km, that quantifies the average contact force in areas of the cell displaying weak bonding. By analogy with classical contact mechanics, we also define the ratio of the real to nominal contact areas, Sr/St. We show that Km can be interpreted as a quantitative indicator of passive contact at metal-cell interfaces, while Sr/St is sensitive to active adhesive processes in the nuclear region. The ability to separate the contributions of passive and active adhesion processes should allow gaining insight into cell-substrate interactions, with important applications in tissue engineering.
15 […]
BERBON, Mélanie; HABENSTEIN, Birgit; LOQUET, Antoine;
Detection of side-chain proton resonances of fully protonated biosolids in nano-litre volumes by magic angle spinning solid-state NMR
Journal of biomolecular NMR, 2018, 0,
We present a new solid-state NMR proton-detected three-dimensional experiment dedicated to the observation of protein proton side chain resonances in nano-liter volumes. The experiment takes advantage of very fast magic angle spinning and double quantum 13C-13C transfer to establish efficient (H)CCH correlations detected on side chain protons. Our approach is demonstrated on the HET-s prion domain in its functional amyloid fibrillar form, fully protonated, with a sample amount of less than 500 g using a MAS frequency of 70kHz. The majority of aliphatic and aromatic side chain protons (70%) are observable, in addition to Halpha resonances, in a single experiment providing a complementary approach to the established proton-detected amide-based multidimensional solid-state NMR experiments for the study and resonance assignment of biosolid samples, in particular for aromatic side chain resonances.
16 […]
KWON, Sunbum; DAWSON, Simon; MANDAL, Pradeep Kumar; HUC, Ivan;
Ribosomal synthesis and folding of peptide-helical aromatic foldamer hybrids
Nature chemistry, 2018, 0,
Translation, the mRNA-templated synthesis of peptides by the ribosome, can be manipulated to incorporate variants of the 20 cognate amino acids. Such approaches for expanding the range of chemical entities that can be produced by the ribosome may accelerate the discovery of molecules that can perform functions for which poorly folded, short peptidic sequences are ill suited. Here, we show that the ribosome tolerates some artificial helical aromatic oligomers, so-called foldamers. Using a flexible tRNA-acylation ribozyme-flexizyme-foldamers were attached to tRNA, and the resulting acylated tRNAs were delivered to the ribosome to initiate the synthesis of non-cyclic and cyclic foldamer-peptide hybrid molecules. Passing through the ribosome exit tunnel requires the foldamers to unfold. Yet foldamers encode sufficient folding information to influence the peptide structure once translation is completed. We also show that in cyclic hybrids, the foldamer portion can fold into a helix and force the peptide segment to adopt a constrained and stretched conformation.
17
SANTARELLI, Xavier;
(FR) COMPOSITIONS DE NANOVECTEURS À BASE DE LIPIDES CHARGÉES DE NANOPARTICULES MÉTALLIQUES ET D'UN AGENT THÉRAPEUTIQUE (EN) LIPID BASED NANOCARRIER COMPOSITIONS LOADED WITH METAL NANOPARTICLES AND THERAPEUTIC AGENT
, 2018, 0,
18 […]
MARTINEZ, Denis; LEGRAND, Anthony; LAMBERT, Olivier; BERBON, Mélanie; GRELARD, Axelle; LOQUET, Antoine; HABENSTEIN, Birgit;
Coiled-coil oligomerization controls localization of the plasma membrane REMORINs
Journal of structural biology, 2018, 0,
REMORINs are nanodomain-organized proteins located in the plasma membrane and involved in cellular responses in plants. The dynamic assembly of the membrane nanodomains represents an essential tool of the versatile membrane barriers to control and modulate cellular functions. Nevertheless, the assembly mechanisms and protein organization strategies of nanodomains are poorly understood and many structural aspects are difficult to visualize. Using an ensemble of biophysical approaches, including solid-state nuclear magnetic resonance, cryo-electron microscopy and in vivo confocal imaging, we provide first insights on the role and the structural mechanisms of REMORIN trimerization. Our results suggest that the formation of REMORIN coiled-coil trimers is essential for membrane recruitment and promotes REMORIN assembly in vitro into long filaments by trimer-trimer interactions that might participate in nanoclustering into membrane domains in vivo.
19 […]
SOLE, Marion; MARIA, Sophie; CABANNE, Charlotte; SANTARELLI, Xavier;
Comparative study of strong cation exchangers: Structure-related chromatographic performances
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2018, 1080, 1-10
Chromatographic performances are highly influenced by operational parameters. New ion exchangers have tailored matrices providing low backpressure, thereby allowing high flow velocity. By systematic frontal analysis and selectivity determination at different flow rates, we independently evaluated cation exchangers to facilitate media selection and investigated the relationship between surface modification and chromatographic performances. Structure-extended resins showed higher binding capacities compared to resins with conventional ligands directly attached to the matrix. Moreover, they maintained high capacities even with high flow velocities. Ligand accessibility was therefore largely enhanced, allowing proteins to interact and bind under harsh conditions with minimal residence/contact time. High throughput resins can be used for purification of high volume and high concentration feedstock in limited time. This results in higher productivity, and could contribute to cost reduction. In this work, we evaluated the dynamic binding capacities of various new ion exchange resins at different binding conductivities for different residence times, and observed that.
20 […]
LECOMTE, Sophie;
Interfacial activity and emulsifying behaviour of inclusion complexes between helical polysaccharides and flavouring molecules resulting from non-covalent interactions
Food Research International, 2018, 105, 801-811
This study deals with the fabrication of inclusion complexes starting from a cross coupling of seven helical polysaccharides (host) and six flavouring agents (guest). Neither of the substrates is considered as an emulsifier when studied alone. Due to a complexation mechanism, the presence of intermolecular hydrogen bonds between substrates was highlighted by infra-red spectroscopy and C-13 NMR. In addition, depending on the polysaccharide used, the guest molecule could be preferentially located either inside or in the interstitial spaces of the helix. In a comparison between raw substrates, the inclusion complexes obtained presented the unique interfacial activity of decreasing surface tension values (gamma) and, in some cases, their behaviour in water was similar to that of regular emulsifiers due to the presence of a critical aggregation concentration (CAC). Substrate concentrations and the ratios between them were the main parameters investigated in this study, which focused on the two inclusion complexes: vanillin/amylose and vanillin/iota-carrageenan. The first decreased gamma values by as much as 53 mN/m with a double transition, whereas the second could cause gamma fall to 36 mN/m with a regular break. In addition, these systems were able to stabilize foams for up to 60 min, which confirmed their unique emulsifying properties.
21 […]
LECOMTE, Sophie;
Tip-Enhanced Raman Spectroscopy: A Tool for Nanoscale Chemical and Structural Characterization of Biomolecules
Chemphyschem, 2018, 19, 8-18
Due to its high molecular sensitivity and spatial optical resolution down to sub-nanometer values, tip-enhanced Raman spectroscopy (TERS) has emerged as a powerful microscopy technique for nanoscale characterization. Progress in TERS instrumentation and in the manufacturing of efficient TERS tips allow for chemical and structural analysis under various experimental conditions (different wavelengths, substrates, and surrounding media). Many biological species have been examined by using this technique. Nucleic acids (individual nucleobases, DNA, and RNA) can show specific TERS features that reveal their composition, conformation, and defects. TERS studies on peptides and proteins (such as amyloid fibrils) provide relevant information on their morphology and structure, leading to valuable insight to their functions and behavior. Finally, lipid layers and membranes, viruses, bacteria, and cells can also be finely characterized. Generalizing TERS measurements in liq- uid medium to study biological systems is the main future challenge.
22 […]
ZIACH, Krzysztof; CHOLLET, Céline; PRABHAKARAN, Panchami; CORVAGLIA, Valentina; BOSE, Partha Pratim; LAXMI-REDDY, Katta; GODDE, Frédéric; SCHMITTER, Jean-Marie; CHAIGNEPAIN, Stéphane; HUC, Ivan;
Single helically folded aromatic oligoamides that mimic the charge surface of double-stranded B-DNA
Nature Chemistry, 2018, 10, 511-518
Numerous essential biomolecular processes require the recognition of DNA surface features by proteins. Molecules mimicking these features could potentially act as decoys and interfere with pharmacologically or therapeutically relevant protein-DNA interactions. Although naturally occurring DNA-mimicking proteins have been described, synthetic tunable molecules that mimic the charge surface of double-stranded DNA are not known. Here, we report the design, synthesis and structural characterization of aromatic oligoamides that fold into single helical conformations and display a double helical array of negatively charged residues in positions that match the phosphate moieties in B-DNA. These molecules were able to inhibit several enzymes possessing non-sequence-selective DNA-binding properties, including topoisomerase 1 and HIV-1 integrase, presumably through specific foldamer-protein interactions, whereas sequence-selective enzymes were not inhibited. Such modular and synthetically accessible DNA mimics provide a versatile platform to design novel inhibitors of protein-DNA interactions.
23 […]
FERRAND, Yann; HUC, Ivan;
Frustrated Helicity: Joining the Diverging Ends of a Stable Aromatic Amide Helix to Form a Fluxional Macrocycle
Angewandte Chemie (International ed. in English), 2018, 0,
Macrocyclization of a stable two-turn helical aromatic pentamide, that is, an object with diverging ends that are not prone to cyclization, was made possible by the transient introduction of disruptors of helicity in the form of acid-labile dimethoxybenzyl tertiary amide substituents. After removal of the helicity disruptors, NMR, X-ray crystallography, and computational studies show that the macrocycle possesses a strained structure that tries to gain as high a helical content as possible despite being cyclic. Two points of disruption of helicity remain, in particular a cis amide bond. This point of disruption of helicity can propagate along the cycle in a fluxional manner according to defined trajectories to produce ten degenerate conformations.
24 […]
HABENSTEIN, Birgit; LOQUET, Antoine;
Femtosecond X-ray coherent diffraction of aligned amyloid fibrils on low background graphene
Nature Communications, 2018, 9,
Here we present a new approach to diffraction imaging of amyloid fibrils, combining a freestanding graphene support and single nanofocused X-ray pulses of femtosecond duration from an X-ray free-electron laser. Due to the very low background scattering from the graphene support and mutual alignment of filaments, diffraction from tobacco mosaic virus (TMV) filaments and amyloid protofibrils is obtained to 2.7 A and 2.4 A resolution in single diffraction patterns, respectively. Some TMV diffraction patterns exhibit asymmetry that indicates the presence of a limited number of axial rotations in the XFEL focus. Signal-to-noise levels from individual diffraction patterns are enhanced using computational alignment and merging, giving patterns that are superior to those obtainable from synchrotron radiation sources. We anticipate that our approach will be a starting point for further investigations into unsolved structures of filaments and other weakly scattering objects.
25 […]
CHADEL, Samy;
CHELATE NANOEMULSION FOR MRI
, 2018, 0,
The present invention relates to an oil-in-water nanoemulsion composition for MRI, including: an aqueous phase which is 70 to 90 wt % of the composition, advantageously 75 to 85 wt %, more advantageously 78 to 82 wt %; a lipid phase including an oil, which is 9.5 to 29.5 wt % of the composition, advantageously 14 to 25 wt %, more advantageously 17 to 21 wt %; a surfactant at the interface between the aqueous and lipid phases, the surfactant including at least one amphiphilic paramagnetic metal chelate and optionally an amphiphilic lipid; the total content by weight of the surfactant relative to the oil being 4 to 10 wt %, advantageously 5 to 8 wt %; the total content by weight of the surfactant relative to the composition being 0.35 to 2.95 wt %, advantageously 0.5 to 2 wt %; the oil including at least 70 wt %, advantageously at least 80 wt %, more advantageously at least 95 wt %, in particular at least 97 wt %, of C6-C18 saturated fatty acids, advantageously C6-C14, and more advantageously C6-C10. (FR)La présente concerne une composition de nanoémulsion huile dans eau pour IRM, comprenant : une phase aqueuse, représentant 70 à 90% en poids de la composition, avantageusement 75 à 85%, plus avantageusement de 78 à 82% une phase lipidique comprenant une huile, représentant 9.5 à 29.5% en poids de la composition, avantageusement 14 à 25%, plus avantageusement 17 à 21%, un tensioactif à l 'interface entre les phases aqueuse et lipidique, le tensioactif comprenant au moins un chélate de métal paramagnétique amphiphile et éventuellement un lipide amphiphile; - la teneur totale en tensioactif en poids par rapport à l'huile étant comprise entre 4 et 10%, avantageusement entre 5 et 8 %; - la teneur totale en tensioactif en poids par rapport à la composition étant comprise entre 0.35 et 2.95%, avantageusement entre 0.5 et 2 %; - l'huile comprenant au moins 70%, avantageusement au moins 80%, de façon avantageuse au moins 95% en poids, notamment au moins 97 %, d'acides gras saturés en C6-C18, avantageusement en C6-C14, plus avantageusement en C6-C10.
26 […]
NASSRA, Merian; BOURGEOIS, Christine; SAUVANT, Patrick; ATGIE, Claude;
Oral administration of lipid oil-in-water emulsions performed with synthetic or protein-type emulsifiers differentially affects post-prandial triacylglycerolemia in rats
Journal of physiology and biochemistry, 2018, 0,
In this study, we compared the impact of administration of size-calibrated lipid emulsions prepared with either synthetic or natural emulsifiers on the post-absorptive plasma triacylglycerol responses in rats. We did this using four types of size-calibrated (10mum diameter) and metastable (3days) emulsions with 20% of an oleic acid-rich sunflower oil and 1% of either synthetic emulsifiers (Tween 80 or sodium 2-stearoyl-lactylate) or two proteins (beta-lactoglobulin or sodium caseinate). An oral fat tolerance test was performed in fasted rats by oral administration of each of these formulations in continuous or emulsified forms. Kinetic parameters (AUC0-inf., AUC0-6h, Cmax, Tmax, and T1/2) for the description of the plasma triacylglycerol responses were calculated. AUC0-6h and AUC0-inf. calculated for the protein groups were significantly lower than those of the control and the synthetic groups. These lower values were associated with significant decreases in the Cmax, exacerbated by the emulsion form and with marked decreases in the Tmax as compared to the control group. T1/2 values were differentially affected by the lipid administration forms and by the nature of the emulsifiers. As compared with the control group, T1/2 was largely increased in the sodium stearoyl-2-lactylate group, but on the contrary, largely lowered in the casein group. We concluded that the use of proteins as natural emulsifiers in lipid emulsions decreased the magnitude of post-prandial triacylglycerolemia for the same amount of ingested lipids, when the emulsion size is controlled for. Proteins could be a promising alternative to the widespread use of synthetic emulsifiers in the food industry.
27 […]
VILAIN, Sébastien;
Growth and Extended Survival of Escherichia coli O157:H7 in Soil Organic Matter
Frontiers in Microbiology, 2018, 9,
Enterohaemorrhagic Escherichia coli, such as serotype O157:H7, are a leading cause of food-associated outbreaks. While the primary reservoir is associated with cattle, plant foods have been associated as sources of human infection. E. coli is able to grow in the tissue of food plants such as spinach. While fecal contamination is the primary suspect, soil has been underestimated as a potential reservoir. Persistence of bacterial populations in open systems is the product of growth, death, predation, and competition. Here we report that E. coli O157:H7 can grow using the soluble compounds in soil, and characterize the effect of soil growth on the stationary phase proteome. E. coli 933D (stxII(-)) was cultured in Soil Extracted Soluble Organic Matter (SESOM) and the culturable count determined for 24d. The proteomes of exponential and stationary phase populations were characterized by 2D gel electrophoresis and protein spots were identified by MALDI-TOF mass spectrometry. While LB controls displayed a death phase, SESOMgrown population remained culturable for 24d, indicating an altered physiological state with superior longevity. This was not due to decreased cell density on entry to stationary phase as 24 h SESOM populations concentrated 10-fold retained their longevity. Principal component analysis showed that stationary phase proteomes from SESOM and LB were different. Differences included proteins involved in stress response, motility, membrane and wall composition, nutrient uptake, translation and protein turnover, and anabolic and catabolic pathways, indicating an altered physiological state of soil-grown cells entering stationary phase. The results suggest that E. coli may be a soil commensal that, in absence of predation and competition, maintains stable populations in soil.
28 […]
MATEUS, Pedro; WICHER, Barbara; FERRAND, Yann; HUC, Ivan;
Carbohydrate binding through first- and second-sphere coordination within aromatic oligoamide metallofoldamers
Chemical communications (Cambridge, England), 2018, 54, 5078-5081
Aromatic oligoamide capsules that fold upon metal binding recognize carbohydrate guests in solution as evidenced by CD and NMR titrations. Crystallographic data reveal that, besides their structural role, metal ions also contribute to guest recognition through either first- or second-sphere coordination.
29 […]
MARTINEZ, Denis; HUGUES, Michel; ODAERT, Benoit;
Assignment of H-1, C-13 and N-15 resonances and secondary structure of the Rgd1-RhoGAP domain
Biomolecular Nmr Assignments, 2018, 12, 129-132
The protein Rgd1 is involved in the regulation of cytoskeleton formation and in signalling pathways that control cell polarity and growth in Saccharomyces cerevisiae. Rgd1p is composed of a F-BAR domain required for membrane binding and a RhoGAP domain responsible for activating Rho3p and Rho4p, two GTPases respectively involved in bud growth and cytokinesis. Rgd1p is recruited to the membrane through interactions with phosphoinositide lipids, which bind the two isolated domains and stimulate the RhoGAP activity on Rho4p. As previously shown by crystallography, the membrane-binding F-BAR domain contains a conserved inositol phosphate binding site, which explains the preferential binding of phosphoinositides. In contrast, RhoGAP domains are not expected to bind lipids. In order to unravel this puzzling feature, we solved the three-dimensional structure of the isolated protein and found a cryptic phosphoinositide binding site involving non conserved residues (Martinez et al. 2017). The assignment of the resonances and secondary structure of Rgd1-RhoGAP (aa 450-666) is presented here.
30 […]
LOQUET, Antoine; BERBON, Mélanie; HABENSTEIN, Birgit;
3D structure determination of amyloid fibrils using solid-state NMR spectroscopy
Methods (San Diego, Calif.), 2018, 0,
The amyloid fold is structurally characterized by a typical cross-beta architecture, which is under debate to represent an energy-favourable folding state that many globular or natively unfolded proteins can adopt. Being initially solely associated with amyloid fibrils observed in the propagation of several neurodegenerative disorders, the discovery of non-pathological (or "functional") amyloids in many native biological processes has recently further intensified the general interest invested in those cross-beta supramolecular assemblies. The insoluble and non-crystalline nature of amyloid fibrils and their usually inhomogeneous appearance on the mesoscopic level pose a challenge to biophysical techniques aiming at an atomic-level structural characterization. Solid-state NMR spectroscopy (SSNMR) has granted breakthroughs in structural investigations on amyloid fibrils ranging from the assessment of the impact of polymorphism in disease development to the 3D atomic structure determination of amyloid fibrils. First landmark studies towards the characterization of atomic structures and interactions involving functional amyloids have provided new impulses in the understanding of the role of the amyloid fold in native biological functions. Over the last decade many strategies have been developed in protein isotope labelling, NMR resonance assignment, distance restraint determination and 3D structure calculation of amyloid fibrils based on SSNMR approaches. We will here discuss the emerging concepts and state-of-the-art methods related to the assessment of amyloid structures and interactions involving amyloid entities by SSNMR.
31 […]
FERRAND, Yann; HUC, Ivan;
Designing Helical Molecular Capsules Based on Folded Aromatic Amide Oligomers
Accounts of Chemical Research, 2018, 51, 970-977
The ab initio rational structure-based design of a synthetic molecular receptor for a given complex biomolecular guest remains an elusive objective, yet remarkable progress has been achieved in recent years. This Account deals with the use of folded artificial aromatic amide oligomers, also termed aromatic foldamers, inspired from biopolymer structures, for the design of helical molecular capsules that can recognize guest molecules, completely surround them, and isolate them from the solvent, thus giving rise to a sort of guest encapsulation associated with slow binding and release kinetics. The development of new amino acid, diacid, and diamine monomers, a main source of creativity in this field, progress in their assembly into ever longer oligoamide sequences, and the predictability of the folded structures due to their inherent rigidity and simple folding principles, allowed for the design and preparation of unimolecular and bimolecular capsule shapes. These capsules consist of molecular helices having a large diameter in the middle and a narrow diameter at both ends thus creating a cavity suitable for binding a guest molecule. The understanding of molecular recognition properties within these bioinspired containers has greatly progressed. Recognition of simple guests such as diols or amino-alcohols may thus be predicted, and hosts can be proposed for guests as complex as saccharides using first principle design. Taking advantage of the modular nature of oligomeric sequences, of their synthetic accessibility and of their propensity to grow into crystals suitable for X-ray crystallographic analysis, a structure-based iterative design methodology has been developed that eventually yielded exquisite guest selectivity, affinity, and diastereoselectivity. This methodology involves rational negative design steps during which changes in the foldamer capsule sequence are not intended to improve binding to the targeted guest but instead to exclude the binding of other guests while preserving key interactions with the target. Metal ions can also be introduced at the inner rim of foldamer capsules and eventually assist the binding of an organic guest. These results demonstrate the viability of an ab initio approach to abiotic receptor design based on aromatic foldamers. The dynamic of the capsules associated with their self-organized nature provides opportunities to not only tune guest binding and selectivity, but also guest capture and release kinetics as well as cavity size and shape. Controlled release thus emerges as a realistic objective. Recent progress thus opens up multiple perspectives for the development of tailored hosts, sensors, and carriers structurally and conceptually different from earlier generations of macrocyclic-based receptors or from supramolecular containers produced by self-assembly.
32 […]
LOMBARDO, Caterina Maria; LECHNER, Marie-Charlotte; GUICHARD, Gilles;
Relationship between the agonist activity of synthetic ligands of TRAIL-R2 and their cell surface binding modes
Oncotarget, 2018, 9, 15566-15578
Tumor Necrosis Factor Receptor Apoptosis Inducing Ligand (TRAIL) appears as an interesting candidate for targeted cancer therapy as it induces apoptosis in cancer cells without toxicity to normal cells. TRAIL elicits apoptosis through agonist death receptor TRAIL-R1 and TRAIL-R2 engagement. Nevertheless, recombinant soluble TRAIL and monoclonal antibodies against these receptors demonstrated insufficient efficacy in clinical trials. This may be explained by the cell-type dependency of the apoptotic response, itself influenced by the effect on ligand binding mode of factors such as the level of receptor oligomerization or glycosylation. To investigate the relation between binding mode and signaling, we used previously described synthetic divalent and monovalent peptides specific for TRAIL-R2. We measured their pro-apoptotic activity on three cancer cell lines sensitive to rhTRAIL induced-apoptosis and monitored their cell-surface binding kinetics. The two divalent peptides bound with strong affinity to TRAIL-R2 expressed on B lymphoma BJAB cells and induced a high degree of apoptosis. By contrast, the same peptides bound weakly to TRAIL-R2 expressed at the surface of the human colon cancer HCT116 or T lymphoma Jurkat cell lines and did not induce their apoptosis. Cross-linking experiments suggest that these differences could be afforded by variations in the TRAIL-R2 oligomerization state at cell surface before ligand addition. Moreover divalent peptides showed a different efficiency in BJAB apoptosis induction, and kinetic distribution analysis of the BJAB binding curves suggested subtle differences in binding mechanisms. Thus our data support a relation between the cell-surface binding mode of the peptides and their pro-apoptotic activity. In this case the precise characterization of ligand binding to the surface of living cells would be predictive of the therapeutic potential of TRAIL-R2 synthetic ligands prior to clinical trials.
33
AZOUZ, Mehdi; CULLIN, Christophe; LECOMTE, Sophie;
Towards a Nanoscale Description of the Interactions between Amyloid Peptide A beta 1-42 and Mutants with Membranes
Biophysical Journal, 2018, 114, 265A-265A
34 […]
PERRIER, Romain; JAFFREDO, Manon; GAITAN, Julien; CATARGI, Bogdan; RAOUX, Matthieu; LANG, Jochen;
Bioelectronic organ-based sensor for microfluidic real-time analysis of the demand in insulin
Biosens Bioelectron, 2018, 117, 253-259
On-line and real-time analysis of micro-organ activity permits to use the endogenous analytical power of cellular signal transduction algorithms as biosensors. We have developed here such a sensor using only a few pancreatic endocrine islets and the avoidance of transgenes or chemical probes reduces bias and procures general usage. Nutrient and hormone-induced changes in islet ion fluxes through channels provide the first integrative read-out of micro-organ activity. Using extracellular electrodes we captured this read-out non-invasively as slow potentials which reflect glucose concentration-dependent (3-15mM) micro-organ activation and coupling. Custom-made PDMS-based microfluidics with platinum black micro-electrode arrays required only some tens of islets and functioned at flow rates of 1-10microl/min which are compatible with microdialysis. We developed hardware solutions for on-line real-time analysis on a reconfigurable Field-Programmable Gate Array (FPGA) that offered resource-efficient architecture and storage of intermediary processing stages. Moreover, real-time adaptive and reconfigurable algorithms accounted for signal disparities and noise distribution. Based on islet slow potentials, this integrated set-up allowed within less than 40mus the discrimination and precise automatic ranking of small increases (2mM steps) of glucose concentrations in real time and within the physiological glucose range. This approach shall permit further development in continuous monitoring of the demand for insulin in type 1 diabetes as well as monitoring of organs-on-chip or maturation of stem-cell derived islets.
35 […]
PERRIER, Romain; RAOUX, Matthieu; JAFFREDO, Manon; LANG, Jochen;
Multimed: An Integrated, Multi-Application Platform for the Real-Time Recording and Sub-Millisecond Processing of Biosignals
Sensors, 2018, 18, 2099
Enhanced understanding and control of electrophysiology mechanisms are increasingly being hailed as key knowledge in the fields of modern biology and medicine. As more and more excitable cell mechanics are being investigated and exploited, the need for flexible electrophysiology setups becomes apparent. With that aim, we designed Multimed, which is a versatile hardware platform for the real-time recording and processing of biosignals. Digital processing in Multimed is an arrangement of generic processing units from a custom library. These can freely be rearranged to match the needs of the application. Embedded onto a Field Programmable Gate Array (FPGA), these modules utilize full-hardware signal processing to lower processing latency. It achieves constant latency, and sub-millisecond processing and decision-making on 64 channels. The FPGA core processing unit makes Multimed suitable as either a reconfigurable electrophysiology system or a prototyping platform for VLSI implantable medical devices. It is specifically designed for open- and closed-loop experiments and provides consistent feedback rules, well within biological microseconds timeframes. This paper presents the specifications and architecture of the Multimed system, then details the biosignal processing algorithms and their digital implementation. Finally, three applications utilizing Multimed in neuroscience and diabetes research are described. They demonstrate the system’s configurability, its multi-channel, real-time processing, and its feedback control capabilities.
36 […]
GAO, Jie; ODA, Reiko; POUGET, Emilie;
Gold Nanoparticle Chains: Synthesis, Characterization, and Modeling Using Spectroscopic Ellipsometry
Journal of Physical Chemistry C, 2018, 122, 11973-11984
In this paper, we explore the ability of ellipsometry to characterize colloidal suspensions composed of gold nanoparticle (NP) chains. The complex effective index of these suspensions is deduced from ellipsometric measurement by using a wavelength-by-wavelength inversion without any dispersion law. We show that the effective refractive index of these colloids is defined by the nature of the solvent, whereas their effective extinction coefficient is mainly sensitive to the plasmonic properties of NP chains. The influence of the NP radius distribution and arrangement on the effective extinction coefficient of NP chain are investigated through simulations based on the coupled point dipole method (CDM). We clearly show that this coefficient is mainly sensitive to the interparticle distance and the number of NPs in the longest segment of chains. We demonstrate that the distribution of the number of NPs in the longest segment of chains and their volume fractions can be directly deduced from the ellipsometry by using the CDM.
37 […]
CARULLA, Natalia;
Direct Evidence of the Presence of Cross-Linked A beta Dimers in the Brains of Alzheimer's Disease Patients
Analytical Chemistry, 2018, 90, 4552-4560
Brain-derived amyloid-beta (A beta) dimers are associated with Alzheimer's disease (AD). However, their covalent nature remains controversial. This feature is relevant, as a covalent cross-link has been proposed to make brain-derived dimers (brain dimers) more synaptotoxic than A beta monomers and would also make them suitable candidates for biomarker development. To resolve this controversy, we here present a three-step approach. First, we validated a type of synthetic cross-linked A beta (CL A beta) dimers, obtained by means of the photoinduced cross-linking of unmodified proteins (PICUP) reaction, as well-defined mimics of putative brain CL A beta dimers. Second, we used these PICUP CL A beta dimers as standards to improve the isolation of brain A beta dimers and to develop state-of-the-art mass spectrometry (MS) strategies to allow their characterization. Third, we applied these MS methods to the analysis of brain A beta dimer samples allowing the detection of the CL [A beta(6-16)](2) peptide comprising a dityrosine cross-link. This result demonstrates the presence of CL A beta dimers in the brains of patients with AD and opens up avenues for establishing new therapeutic targets and developing novel biomarkers for this disease.
38 […]
BRESSOLLIER, Philippe; URDACI, Maria;
Lactobacillus rhamnosus GG increases COX-2 expression and PGE2 secretion in colonic myofibroblasts via a MyD88-dependent mechanism during homeostasis
Cellular microbiology, 2018, 0, e12871-e12871
Prostaglandin E2 (PGE2 ) plays a critical role in intestinal mucosal tolerance and barrier integrity. Cyclooxygenase-2 (COX-2)-dependent PGE2 production involves mobilization of arachidonic acid (AA). Lactobacillus rhamnosus GG (LbGG) is one of the most widely used probiotics reported to colonize the colonic mucosa. LbGG contributes to the protection of the small intestine against radiation injury through the repositioning of mucosal COX-2 expressing cells. However, it is unknown if LbGG modulates PGE2 production in the colonic mucosa under homeostasis and the major cellular elements involved in these processes. Colonic epithelial and CD90+ mesenchymal stromal cells, also known as (myo) fibroblasts (CMFs), are abundant innate immune cells in normal colonic mucosa able to produce PGE2 . Herein, we tested the hypothesis that under colonic mucosal homeostasis LbGG modulates the eicosanoid pathway resulting in increased PGE2 production in both epithelial and stromal cells. Among the five tested human colonic epithelial cell lines, only exposure of Caco-2 to LbGG for 24 h led to the mobilization of arachidonic acid (AA) with concomitant increase in the components within the leukotriene and COX-2 dependent PGE2 pathways. By contrast, CMFs isolated from the normal human colonic mucosa responded to LbGG with increased expression of COX-2 and PGE2 in the prostaglandin pathway, but not 5-LO in the leukotriene pathway. Oral gavage of C57BL/6 mice for 5 days with LbGG (5x108 CFU/dose) increased COX-2 expression in the colonic mucosa. The majority of cells upregulating COX-2 protein expression were located in the colonic lamina propria, and co-localized with alpha-SMA+ cells corresponding to the CMF phenotype. This process was MyD88 dependent, since silencing of MyD88 expression in CMFs abrogated LbGG-induced upregulation of COX-2 in culture and in vivo. Taken together, our data suggests that LbGG increases release of COX-2 mediated PGE2 , contributing to the maintenance of mucosal homeostasis in the colon and CMFs are among the major contributors to this process.
39 […]
CARULLA, Natalia;
Stabilization of a Membrane-Associated Amyloid-beta Oligomer for Its Validation in Alzheimer's Disease
Frontiers in molecular biosciences, 2018, 5, 38-38
We have recently reported on the preparation of a membrane-associated beta-barrel Pore-Forming Abeta42 Oligomer (betaPFOAbeta42). It corresponds to a stable and homogeneous Abeta42 oligomer that inserts into lipid bilayers as a well-defined pore and adopts a specific structure with characteristics of a beta-barrel arrangement. As a follow-up of this work, we aim to establish betaPFOAbeta42's relevance in Alzheimer's disease (AD). However, betaPFOAbeta42 is formed under dodecyl phosphocholine (DPC) micelle conditions-intended to mimic the hydrophobic environment of membranes-which are dynamic. Consequently, dilution of the betaPFOAbeta42/DPC complex in a detergent-free buffer leads to dispersion of the DPC molecules from the oligomer surface, leaving the oligomer without the hydrophobic micelle belt that stabilizes it. Since dilution is required for any biological test, transfer of betaPFOAbeta42 from DPC micelles into another hydrophobic biomimetic membrane environment, that remains associated with betaPFOAbeta42 even under high dilution conditions, is a requisite for the validation of betaPFOAbeta42 in AD. Here we describe conditions for exchanging DPC micelles with amphipols (APols), which are amphipathic polymers designed to stabilize membrane proteins in aqueous solutions. APols bind in an irreversible but non-covalent manner to the hydrophobic surface of membrane proteins preserving their structure even under extreme dilution conditions. We tested three types of APols with distinct physical-chemical properties and found that the betaPFOAbeta42/DPC complex can only be trapped in non-ionic APols (NAPols). The characterization of the resulting betaPFOAbeta42/NAPol complex by biochemical tools and structural biology techniques allowed us to establish that the oligomer structure is maintained even under high dilution. Based on these findings, this work constitutes a first step towards the in vivo validation of betaPFOAbeta42 in AD.
40 […]
NLATE, Sylvain; ATTOUI, Mariam; POUGET, Emilie; ODA, Reiko;
Optically Active Polyoxometalate-based Silica Nanohelices: Induced Chirality from Inorganic Nanohelices to Achiral POM Clusters
Chemistry (Weinheim an der Bergstrasse, Germany), 2018, 0,
In order to investigate the principle of chiral induction from nanometric silica helices to polyoxometalate (POM) clusters, a series of optically active silica POM-based nanohelices (NANOPOMs) have been prepared by electrostatic grafting and direct adsorption of -Keggin polyoxometalate [-PW12O40]3- to well-defined left- and right-handed silica nanohelices. UV-vis, Raman, DRIFT, TEM, HR-TEM, EDS and circular dichroism (CD) spectroscopy were used to characterize these NANOPOMs, and confirm the presence of POM clusters as well as their interactions with the helical support. The optical activity of the left-handed and right-handed NANOPOMs has been proven by CD spectroscopy. Their CD spectra are mirror images of one another, showing cotton effects at around 214 and 276 nm, this last contribution corresponding to the oxygen-to-tungsten charge-transfer bands of Keggin polyoxoanions. The CD signal of POM clusters is strongly enhanced for NANOPOMs built by adsorption of POM onto silica nanohelices, indicating a better induced optical activity to POM clusters. These nanohelices are stable, recoverable and active catalysts in the oxidation of sulfides. To the best of our knowledge, the present research represents the first examples of optically active POM-containing silica nanohelices in which achiral POM clusters have been grafted onto silica nanohelices, and display chiroptical effects.
41 […]
BONNEU, Marc;
Energy Metabolism Rewiring Precedes UVB-Induced Primary Skin Tumor Formation
Cell Reports, 2018, 23, 3621-3634
Although growing evidence indicates that bioenergetic metabolism plays an important role in the progression of tumorigenesis, little information is available on the contribution of reprogramming of energy metabolism in cancer initiation. By applying a quantitative proteomic approach and targeted metabolomics, we find that specific metabolic modifications precede primary skin tumor formation. Using a multistage model of ultraviolet B (UVB) radiation-induced skin cancer, we show that glycolysis, tricarboxylic acid (TCA) cycle, and fatty acid beta-oxidation are decreased at a very early stage of photocarcinogenesis, while the distal part of the electron transport chain (ETC) is upregulated. Reductive glutamine metabolism and the activity of dihydroorotate dehydrogenase (DHODH) are both necessary for maintaining high ETC. Mice with decreased DHODH activity or impaired ETC failed to develop pre-malignant and malignant lesions. DHODH activity represents a major link between DNA repair efficiency and bioenergetic patterning during skin carcinogenesis.
42 […]
CIACCAFAVA, Alexandre;
Controlling Redox Enzyme Orientation at Planar Electrodes
Catalysts, 2018, 8,
Redox enzymes, which catalyze reactions involving electron transfers in living organisms, are very promising components of biotechnological devices, and can be envisioned for sensing applications as well as for energy conversion. In this context, one of the most significant challenges is to achieve efficient direct electron transfer by tunneling between enzymes and conductive surfaces. Based on various examples of bioelectrochemical studies described in the recent literature, this review discusses the issue of enzyme immobilization at planar electrode interfaces. The fundamental importance of controlling enzyme orientation, how to obtain such orientation, and how it can be verified experimentally or by modeling are the three main directions explored. Since redox enzymes are sizable proteins with anisotropic properties, achieving their functional immobilization requires a specific and controlled orientation on the electrode surface. All the factors influenced by this orientation are described, ranging from electronic conductivity to efficiency of substrate supply. The specificities of the enzymatic molecule, surface properties, and dipole moment, which in turn influence the orientation, are introduced. Various ways of ensuring functional immobilization through tuning of both the enzyme and the electrode surface are then described. Finally, the review deals with analytical techniques that have enabled characterization and quantification of successful achievement of the desired orientation. The rich contributions of electrochemistry, spectroscopy (especially infrared spectroscopy), modeling, and microscopy are featured, along with their limitations.
43 […]
DUFFAU, Emilie; HARTE, Etienne; LECOMTE, Sophie; LEAL-CALDERON, Fernando; FAURE, Chrystel;
Influence of Implants Composition on Melatonin Release from Ethylcellulose Matrix
Current Drug Delivery, 2018, 15, 737-743
Background: Melatonin release from Ethylcellulose matrix has never been studied on the whole range of compositions. Objective: To perform a comprehensive study about the influence of the melatonin loading on its release from solid ethylcellulose implants, from both a kinetic and structural point of view. Method: Cylindrical implants differing in their Melatonin:Ethylcellulose ratio were fabricated to cover a large range of compositions. Drug release was assayed by in vitro dissolution tests in CTAB micellar solutions. The 2D imaging of implant chemical composition during Melatonin release was performed by confocal Raman spectroscopy. FT-IR spectroscopy and Karl-Fisher technique were employed to study implants hydration. Results: A drug radial leakage, whatever the implant composition, is imaged. The apparent diffusion coefficient, D of melatonin was evaluated considering Fickian radial diffusion: its value ranges from 2 to 6 10(-12) cm(2)/s depending on the EC content. The variation of the characteristic drug delivery time with composition was non-monotonous and two different regimes were identified. Conclusion: A micellar transport of Melatonin was found. The two regimes in drug release were interpreted considering the polymer barrier effect, the initial porosity and M domains connectivity.
44 […]
DRIDI, Wafa; MONTEIL, Julien; FAURE, Chrystel; LEAL-CALDERON, Fernando;
Monodisperse O/W emulsions stabilized by proteins: how to master the average droplet size and stability, while minimizing the amount of proteins
Langmuir : the ACS journal of surfaces and colloids, 2018, 0,
Hexadecane-in-water emulsions were fabricated by means of a microfluidizer using two types of protein stabilizers, sodium caseinate (NaCAS) and beta-lactoglobulin (BLG). A study of the dependence of the mean droplet diameter and protein coverage on protein concentration was performed. At low protein concentrations, the emulsions were monodisperse and their mean droplet size was governed by the so-called limited-coalescence process. In this regime, the interfacial coverage was constant and was deduced from the linear evolution of the total interfacial area as a function of the amount of adsorbed proteins. In emulsions based on NaCAS, almost all the initial protein content was adsorbed at the interfaces. Emulsions formulated at very low protein content underwent unlimited coalescence after prolonged storage or when submitted to centrifugation. Additional NaCAS was incorporated in the continuous phase, right after the emulsification process, as a means of ensuring kinetic stability. The interfacial coverage increased after protein addition. Other strategies including acidification and salt addition were also probed to gain stability. Instead, in emulsions based on BLG, only partial adsorption of the initial protein content was observed. The corresponding emulsions remained kinetically stable against coalescence and no further addition of protein was required after emulsification. Our approach allows to obtain monodisperse, kinetically stable emulsions, and to master their average droplet size, while minimizing the amount of proteins.
45 […]
BAUDIN, Antoine; COLLIE, Gavin; CHAIGNEPAIN, Stéphane; HOCQUELLET, Agnès; BERBON, Mélanie; LOQUET, Antoine; GUICHARD, Gilles; ODAERT, Benoit;
1H, 13C, 15N NMR resonance assignments and secondary structure determination of the extra-cellular domain from the human proapoptotic TRAIL-R2 death receptor 5 (DR5-ECD)
Biomolecular NMR assignments, 2018, 0,
Death receptors (DR) selectively drive cancer cells to apoptosis upon binding to the Tumor necrosis factor-a-Related Apoptosis-Inducing Ligand (TRAIL). Complex formation induces the oligomerization of the death receptors DR4 (TRAIL-R1) and DR5 (TRAIL-R2) and transduces the apoptogenic signal to their respective death domains, leading to Death Inducing Signaling Complex (DISC) formation, caspase activation and ultimately cell death. Several crystal structures of the ExtraCellular Domain from Death Receptor 5 (DR5-ECD) have been reported in complex with the TRAIL ligand or anti-DR5 antibodies, but none for the isolated protein. In order to fill this gap and to perform binding experiments with TRAIL peptidomimetics, we have produced isotopically labelled DR5-ECD and started a conformational analysis by using high-field 3D NMR spectroscopy. Herein, we present the first resonance assignment of a TRAIL receptor in solution and the determination of its secondary structure from NMR chemical shifts.
Groupe de Bernard Gallois Accueil Members Publications
informations Dernière mise à jour : 20/03/2014


CNRS Université de Bordeaux Institut Polytechnique de Bordeaux