Les bicelles biphényles : un nouveau modèle de biomembrane pour l'étude de protéines membranaires par RMN des solides.

Nous avons développé un nouveau modèle de biomembrane de type bicelle, dont la normale n, s'oriente parallèlement au champ magnétique B0, sans ajout d'ions paramagnétiques. Ces nouveaux objets, de forme bicouche discoïdale, ont pu être observés par microscopie électronique et un diamètre moyen de 800 Å a été statistiquement déterminé. Ce système est constitué d'un mélange approprié de lipides à chaînes courtes (DCPC) et à chaînes longues, le 1-Tétradécanoyl-2-(4-(4-Biphényl)Butanoyl)-sn-glycero-3-PhosphatidylCholine (TBBPC). La présence de deux cycles phényles sur l'une des chaînes aliphatiques confère au TBBPC une anisotropie de susceptibilité magnétique positive, à l'origine de l'orientation spécifique n//B0 des bicelles TBBPC/DCPC. Cette orientation a été caractérisée par diffusion des rayons X aux petits angles et par RMN des solides 31P, 2H et 14N. Les domaines déxistence de ces bicelles en fonction de la composition lipidique, de la température et de l'hydratation, ont ainsi pu être déterminés et comparés à ceux du système DMPC/DCPC, dont la normale s'oriente perpendiculairement à B0. Par l'analyse RMN-2H du TBBPC-2H27, nous avons évalué la dynamique des bicelles biphényles, qui est équivalente à celle des bicelles saturées. Nous avons également montré par diffusion des rayons X, que ces bicelles TBBPC/DCPC conservent leur orientation spécifique n//B0 pendant plusieurs jours en dehors du champ magnétique, ce qui peut trouver de nombreuses applications dans des études biophysiques. Enfin, ces nouveaux modèles membranaires présentent des avantages pour l'étude structurale et orientationnelle (par RMN-15N) de biomolécules telles que des peptides amphipatiques.We have developed a new type of bicelles, which orient such as their normal n aligns parallel to the magnetic field B0 without paramagnetic ions. These new discoidal nano-objects were observed by freeze fracture electron microscopy and an average diameter of 800 Å was statistically determined. This system is composed of an appropriate mixture of short chain (DCPC) and long chain lipids (1-Tetradecanoyl-2-(4-(4-Biphenyl)Butanoyl)-sn-glycero-3-PhosphatidylCholine, TBBPC). The presence of two phenyl rings on one of the aliphatic chains of TBBPC confers to this molecule a positive anisotropic susceptibility, such that TBBPC/DCPC bicelles orient with their normal parallel to B0. This specific orientation was characterized by SAXS analysis and by 31P, 2H and 14N solid state NMR. Consequently, partial temperature-composition-hydration diagrams of these new bicelles were established by 31P-NMR and compared to the ones of DMPC/DCPC bicelles, which orient with their normal perpendicular to B0. The dynamics of biphenyl bicelles was probed by 2H NMR, analysing the TBBPC-2H27, and is equivalent to the one of saturated bicelles. We also proved by X-ray scattering that TBBPC/DCPC bicelles can keep their specific orientation n//B0 outside the magnetic field for several days, which can be useful for biophysical applications. Finally, these new bicelles are shown to be very promising to study the structure and the orientation (by 15N-NMR)of biomolecules such as amphipathic peptides

Les bicelles biphényles (un nouveau modèle de biomenbrane pour l'étude de protéines membranaires par RMN des solides)

Nous avons développé un nouveau modèle de biomembrane de type bicelle, dont la normale n, s oriente parallèlement au champ magnétique B0, sans ajout d ions paramagnétiques. Ces nouveaux objets, de forme bicouche discoïdale, ont pu être observés par microscopie électronique et un diamètre moyen de 800 Å a été statistiquement déterminé. Ce système est constitué d'un mélange approprié de lipides à chaînes courtes (DCPC) et à chaînes longues, le 1-Tétradécanoyl-2-(4-(4-Biphényl)Butanoyl)-sn-glycero-3-PhosphatidylCholine (TBBPC). La présence de deux cycles phényles sur l une des chaînes aliphatiques confère au TBBPC une anisotropie de susceptibilité magnétique positive, à l origine de l orientation spécifique n//B0 des bicelles TBBPC/DCPC. Cette orientation a été caractérisée par diffusion des rayons X aux petits angles et par RMN des solides 31P, 2H et 14N. Les domaines d existence de ces bicelles en fonction de la composition lipidique, de la température et de l hydratation, ont ainsi pu être déterminés et comparés à ceux du système DMPC/DCPC, dont la normale s oriente perpendiculairement à B0. Par l analyse RMN-2H du TBBPC-2H27, nous avons évalué la dynamique des bicelles biphényles, qui est équivalente à celle des bicelles saturées. Nous avons également montré par diffusion des rayons X, que ces bicelles TBBPC/DCPC conservent leur orientation spécifique n//B0 pendant plusieurs jours en dehors du champ magnétique, ce qui peut trouver de nombreuses applications dans des études biophysiques. Enfin, ces nouveaux modèles membranaires présentent des avantages pour l étude structurale et orientationnelle (par RMN-15N) de biomolécules telles que des peptides amphipatiques.BORDEAUX1-BU Sciences-Talence (335222101) / SudocSudocFranceF

Les liposomes biphényles (caractérisation par RMN des solides, microscopies optiques et électroniques et SAXS)

Un nouveau modèle de biomembrane de type liposome a été développé à partir de lipides synthétisés comportant une unité biphényle sur leur chaînes sn2 et une chaîne aliphatique sn1 de longueur et insaturation variables. L anisotropie de susceptibilité magnétique positive de ces molécules induit une déformation en oblate de ces liposomes dits biphényles dans le champ magnétique B0. Cette déformation spécifique a été caractérisée par RMN des solides 31P et 2H en faisant varier différents paramètres : l intensité de B0, l'élasticité membranaire, la température et la taille des liposomes (Helfrich, 1973). Ces vésicules déformées ont pu être observées par microscopies optiques et électroniques et la rémanence de la déformation en dehors de B0 a pu être analysée par diffusion des rayons X aux petits angles (SAXS). Enfin, les premières applications des liposomes biphényles comme nouveau modèle de biomembrane pour analyser la structure et l orientation (par RMN des solides 15N) de peptides ou protéines membranaires, ont été étudiées.A new model of biomembrane (liposome) was developed from synthesized lipids containing a biphenyl unit on the sn2 aliphatic chain and possessing a sn1 aliphatic chain which varies in length and unsaturation. The positive magnetic susceptibility anisotropy of these molecules induces an oblate deformation of these biphenyls liposomes under the magnetic field B0. This particular deformation has been characterized by 31P and 2H solid state NMR by varying different parameters: the intensity of B0, the membrane elasticity, the temperature and the size of the liposomes (Helfrich, 1973). These deformed vesicles were observed by optical and electron microscopy and the remanence of the deformation outside B0 has been analyzed by Small angles X-ray scattering (SAXS).Finally, the first applications of biphenyls liposomes as new biomembrane model to analyze the structure and orientation of membrane proteins or peptides were studied by 15N solid state NMRBORDEAUX1-Bib.electronique (335229901) / SudocSudocFranceF

Observer la biodiversité fonctionnelle en arboriculture : aller jusqu'à l'évaluation

This video presents three monitoring methods (threshing, trap strip and predation plate) that can be used in orchards to monitor functional biodiversity and assess the presence or activity of useful beneficials to control certain pests. The advantage of these simplified methods is that fruit farmers can use them independently to assess the effect of their agro-ecological practices or infrastructures on functional biodiversity

Unprecedented Observation of Days-Long Remnant Orientation of Phospholipid Bicelles: A Small-Angle X-ray Scattering and Theoretical Study

Nanometric bilayer-based self-assembled micelles commonly named as bicelles, formed with a mixture of long and short chains phosphatidylcholine lipids (PC), are known to orient spontaneously in a magnetic field. This field-induced orientational order strongly depends on the molecular structure of the phospholipids. Using small-angle X-ray scattering (SAXS), we performed detailed structural studies of bicelles and investigated the orientation/relaxation kinetics in three different systems: saturated-chain lipid bicelles made of DMPC (dimyristoyl PC)/DCPC (1,2-dicaproyl PC) with and without the added paramagnetic lanthanide ions Eu3+, as well as bicelles of TBBPC (1-tetradecanoyl- 2-(4-(4 biphenyl)butanoyl)-sn-glycero-3-PC)/DCPC. The structural study confirmed the previous NMR studies, which showed that DMPC bicelles orient with the membrane normal perpendicular (defined here as "nematic" orientation) to the magnetic field, whereas they orient parallel (defined here as "smectic" orientation) to the magnetic field in the presence of Eu3+. The TBBPC bicelles also show smectic orientation. Surprisingly, the orientational order induced in the magnetic field remains even after the magnetic field is removed, which allowed us to investigate the orientation and relaxation kinetics of different bicelle structures. We demonstrate that this kinetics is very different for all three types of bicelles at the same lipid concentration; DMPC bicelles (∼40 nm diameter) with and without Eu3+ orient faster than TBBPC bicelles (∼80 nm diameter).However, for the relaxation, DMPC bicelles (nematic) lose their macroscopic orientation only after one hour, whereas both DMPC bicelles with Eu3+ and TBBPC bicelles (smectic) remarkably stay oriented for up to several days! These results indicate that the orientation mechanism of these nanometric disks in the magnetic field is governed by their size, with smaller bicelles orienting faster than the larger bicelles. Their relaxation mechanism outside the magnetic field, however, is governed by the degree of ordering. Indeed, the angular distribution of oriented bicelles is much narrower for the bicelles with smectic orientation, and, consequently, they keep aligned for much longer time (days) than those with nematic ordering (hours) outside the magnetic field. The understanding of the orientation/relaxation kinetics, as well as the morphologies of these "molecular goniometers" at molecular and supramolecular levels, allows controlling such an unprecedented long-range and long-lived smectic ordering of nanodisks and opens a wide field of applications for structural biology or material sciences

Monitoring functional agrobiodiversity in orchards : Going to Assessment !

This video presents three monitoring methods (threshing, trap strip and predation plate) that can be used in orchards to monitor functional biodiversity and assess the presence or activity of useful beneficials to control certain pests. The advantage of these simplified methods is that fruit farmers can use them independently to assess the effect of their agro-ecological practices or infrastructures on functional biodiversity

Natural variation at the FRD3 MATE transporter locus reveals cross-talk between Fe homeostasis and Zn tolerance in Arabidopsis thaliana

Zinc (Zn) is essential for the optimal growth of plants but is toxic if present in excess, so Zn homeostasis needs to be finely tuned. Understanding Zn homeostasis mechanisms in plants will help in the development of innovative approaches for the phytoremediation of Zn-contaminated sites. In this study, Zn tolerance quantitative trait loci (QTL) were identified by analyzing differences in the Bay-0 and Shahdara accessions of Arabidopsis thaliana. Fine-scale mapping showed that a variant of the Fe homeostasis-related FERRIC REDUCTASE DEFECTIVE3 (FRD3) gene, which encodes a multidrug and toxin efflux (MATE) transporter, is responsible for reduced Zn tolerance in A. thaliana. Allelic variation in FRD3 revealed which amino acids are necessary for FRD3 function. In addition, the results of allele-specific expression assays in F1 individuals provide evidence for the existence of at least one putative metal-responsive cis-regulatory element. Our results suggest that FRD3 works as a multimer and is involved in loading Zn into xylem. Cross-homeostasis between Fe and Zn therefore appears to be important for Zn tolerance in A. thaliana with FRD3 acting as an essential regulator

Natural Variation in the ATPS1 Isoform of ATP Sulfurylase Contributes to the Control of Sulfate Levels in Arabidopsis

Sulfur is an essential macronutrient for all living organisms. Plants take up inorganic sulfate from the soil, reduce it, and assimilate it into bioorganic compounds, but part of this sulfate is stored in the vacuoles. In our first attempt to identify genes involved in the control of sulfate content in the leaves, we reported that a quantitative trait locus (QTL) for sulfate content in Arabidopsis (Arabidopsis thaliana) was underlain by the APR2 isoform of the key enzyme of sulfate assimilation, adenosine 5'-phosphosulfate reductase. To increase the knowledge of the control of this trait, we cloned a second QTL from the same analysis. Surprisingly, the gene underlying this QTL encodes the ATPS1 isoform of the enzyme ATP sulfurylase, which precedes adenosine 5'-phosphosulfate reductase in the sulfate assimilation pathway. Plants with the Bay allele of ATPS1 accumulate lower steady-state levels of ATPS1 transcript than those with the Sha allele, which leads to lower enzyme activity and, ultimately, the accumulation of sulfate. Our results show that the transcript variation is controlled in cis. Examination of ATPS1 sequences of Bay-0 and Shahdara identified two deletions in the first intron and immediately downstream the gene in Bay-0 shared with multiple other Arabidopsis accessions. The average ATPS1 transcript levels are lower in these accessions than in those without the deletions, while sulfate levels are significantly higher. Thus, sulfate content in Arabidopsis is controlled by two genes encoding subsequent enzymes in the sulfate assimilation pathway but using different mechanisms, variation in amino acid sequence and variation in expression levels