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1
DITERLIZZI, MARIANNA. « Polymeric Water-Processable Nanoparticles towards sustainable organic photovoltaics ». Doctoral thesis, Università degli Studi di Milano-Bicocca, 2022. http://hdl.handle.net/10281/376407.
Texte intégralRésumé :
Il mio progetto di dottorato è focalizzato sullo sviluppo di inchiostri a base acquosa costituiti da nanoparticelle (NPs) polimeriche per applicazioni optoelettroniche ed elettroniche. In particolare, lo scopo della mia ricerca è la preparazione di strati attivi di dispositivi fotovoltaici organici (OPV) sostenibili. Questo obiettivo è raggiunto attraverso sospensioni acquose di NPs processabili in acqua, preparate a partire da miscele di polimeri semiconduttori donatori e accettori di elettroni. Gli inchiostri acquosi sono stati ottenuti attraverso il metodo della miniemulsione, modificato affinché non fosse necessaria l'aggiunta di surfattanti per garantire la stabilità colloidale. L'approccio sviluppato prevede l'utilizzo di copolimeri a blocchi (BCPs) anfifilici del tipo rod-coil, caratterizzati da un blocco rigido (un polimero semiconduttore di tipo p) legato covalentemente a un segmento flessibile idrofilo in grado di interagire con il mezzo acquoso, stabilizzando le interfacce acquose/non acquose. I BCPs anfifilici sono in grado di auto-assemblarsi sia puri che in miscela con materiali accettori, portando alla formazione di nanostrutture costituite da domini con dimensioni adatte alla percolazione della carica nello strato attivo della cella solare organica (OSC).
In primo luogo, come materiali elettron-donatori sono stati considerati dei polimeri low band-gap (LBG). Nella parte iniziale della tesi è descritta la sintesi di quattro BCPs basati sul polimero poli[2,6-(4,4-bis-(2-etilesil)-4H-ciclopenta[2,1-b;3,4-b']ditiofene)-alt-4,7(2,1,3-benzotiazolo)] (PCPDTBT), con un segmento di poli-4-vinilpiridina (P4VP) come coil. I BCPs sono stati utilizzati in miscela con il derivato fullerenico PC61BM come materiale elettron-accettore per ottenere inchiostri acquosi, che sono stati poi depositati per fabbricare strati attivi. In seguito, sono stati condotti diversi esperimenti per trovare la correlazione tra la morfologia interna e la composizione delle NPs con l'efficienza dei dispositivi OPV.
Successivamente, sono stati studiati altri polimeri LBG dotati di un parziale grado di cristallinità, al fine di migliorare l'efficacia del metodo sviluppato. Pertanto, nella seconda parte della tesi si discute la sintesi e la caratterizzazione di un nuovo BCP anfifilico basato sul poli[[4,8-bis[(2-etilesil)ossi]benzo[1,2-b:4,5-b']ditiofene-2,6-diil][3-fluoro-2-[(2-etilesil)carbonil]tieno[3,4-b]tiofenediil]] (PTB7) come blocco rigido, che è più rigido e più cristallino del PCPDTBT. Come blocco coil è stato scelto un segmento costituito da 15 unità di 4VP. Quindi sono state preparate le NPs tramite self-assembly del PTB7-b-P4VP miscelato con il derivato fullerenico PC71BM. Le sospensioni acquose ottenute sono state impiegate per fabbricare dispositivi OPV in configurazione diretta, e la cella migliore che è stata ottenuta presenta un’efficienza pari a 0.85%, che è un valore ancora molto lontano dal benchmark, ma è comunque superiore all'efficienza del dispositivo ottenuto depositando la miscela PC71BM:PTB7-b-P4VP da solventi alogenati.
Infine, è stato preso in considerazione l'uso di tensioattivi nella preparazione delle NPs, in quanto le sospensioni acquose che ne risultano sono più stabili e più facili da maneggiare e conservare, facilitando il processo di scale-up a livello industriale. In quest’ultima parte della tesi, sono stati studiati altri polimeri semiconduttori come materiali elettron-donatori. In particolare, sono stati sintetizzati e caratterizzati due nuovi polimeri semiconduttori LBG e uno a medio band-gap. Questi materiali saranno miscelati con accettori fullerenici e non per ottenere inchiostri a base acquosa che saranno depositati come strati attivi di dispositivi optoelettronici, analogamente a quanto fatto per i materiali precedenti.My PhD project is focused on the development of polymeric nanoparticle-based aqueous inks for optoelectronic and electronic applications. Specifically, the aim of my research is the fabrication of sustainable active layers of organic photovoltaic (OPV) devices processable in water. This goal is reached through water-processable nanoparticle (WPNP) aqueous suspensions, prepared from semiconducting polymers as electron-donor and acceptor materials. The aqueous inks are obtained through a modified miniemulsion method, which unlike the standard process does not imply the addition of any surfactant to ensure the colloidal stability. The adapted approach involves the use of amphiphilic rod-coil block copolymers (BCPs), characterized by a rigid block (a p‐type semiconducting polymer) covalently linked to a hydrophilic flexible segment able to interact with aqueous medium, stabilizing the aqueous/non-aqueous interfaces. The amphiphilic BCPs are able to self-assemble both neat and in blend with acceptor materials, leading to the formation of nanostructures consisting of domains with dimensions suitable for the charge percolation in the resulting active layer of the organic solar cell (OSC). Primarily, low-band-gap (LBG) polymers were considered as electron donor materials to match the solar radiation absorption. Firstly, the synthesis of four different poly[2,6-(4,4-bis-(2-ethylhexyl)-4H-cyclopenta[2,1-b;3,4-b’]dithiophene)-alt-4,7(2,1,3-benzothiadiazole)] (PCPDTBT)-based amphiphilic BCPs, with a tailored segment of poly-4-vinylpiridine (P4VP) as coil, was presented. The BCPs were used in blend with the [6,6]-phenyl-C61-butyric acid methyl ester (PC61BM) as acceptor material to prepare WPNP aqueous inks, which were deposited to obtain the active layers. The correlation between the internal morphology and composition of the WPNPs, and the dimensions of the donor/acceptor nanodomains with the efficiency of the resulting OSCs was deeply studied. In a second time, we explored other LBG polymers endowed with a partial order to improve the effectiveness of the approach. Therefore, the synthesis and the deep characterization of a new amphiphilic BCP based on the poly[[4,8-bis[(2-ethylhexyl)oxy]benzo[1,2-b:4,5-b’]dithiophene-2,6-diyl][3-fluoro-2-[(2-ethylhexyl)carbonyl]thieno[3,4-b]thiophenediyl]] (PTB7) as rigid donor polymer, which is stiffer and more crystalline than PCPDTBT, were described. A segment of 15 repeating units of 4VP was selected as coil. We prepared WPNPs coming from the self-assembly of the PTB7-b-P4VP blended with the [6,6]-phenyl-C71-butyric acid methyl ester (PC71BM). Subsequently, the WPNPs were employed to fabricate OSCs in direct configuration, and the best gained OPV device exhibited a PCE of 0.85%, which is still very far from the benchmark, but it is higher than the efficiency of the device obtained depositing the PC71BM:PTB7-b-P4VP from halogenated solvents. Lastly, the use of surfactants in the WPNP preparation was considered, as the resulting aqueous suspensions are more stable and easier to handle and store, enhancing the industrial scale-up process. Other semiconducting polymers were selected as electron-donor materials in the active blends. Particularly, two new LBG semiconducting BDT-based polymers, and a medium band-gap one, were synthetized and characterized. These materials will be blended with fullerene and non-fullerene acceptor (NFA) materials to obtain aqueous inks that will be deposited as active layers of optoelectronic devices, similarly to previous materials.
2
Imai, Tomoya. « Nanodomain Structure of Native Cellilose Microfibril ». Kyoto University, 2000. http://hdl.handle.net/2433/78103.
Texte intégralRésumé :
Kyoto University (京都大学)0048
新制・課程博士
博士(農学)
甲第8426号
農博第1110号
新制||農||800(附属図書館)
学位論文||H12||N3383(農学部図書室)
UT51-2000-F330
京都大学大学院農学研究科森林科学専攻
(主査)教授 伊東 隆夫, 教授 東 順一, 教授 藤田 稔
学位規則第4条第1項該当
3
Legrand, Anthony. « Anchoring mechanism of the plant protein remorin to membrane nanodomains ». Thesis, Bordeaux, 2020. http://www.theses.fr/2020BORD0285.
Texte intégralRésumé :
La rémorine du groupe 1 isoforme 3 de Solanum tuberosum (StREM1.3) est une protéine membranaire de la famille multigénique de protéines de plante appelée rémorines (REMs), impliquées dans l’immunité des plantes, la symbiose, la résistance aux stress abiotiques et la signalisation hormonale. La caractéristique la plus connue des REMs est leur capacité à se ségréger en nanodomaines au feuillet interne de la membrane plasmique (MP). Pour StREM1.3, ceci se fait via une interaction entre deux lysines de l’ancre C-terminale de la rémorine (RemCA) et le phosphatidylinositol 4-phosphate (PI4P) négativement chargé. Ainsi, RemCA modifie sa conformation et s’enfonce partiellement dans la MP, résultant en un accrochage membranaire intrinsèque. Capitalisant sur les données structurales déjà disponibles concernant cet isoforme, nous investiguons StREM1.3 davantage quant à ses propriétés d’interaction membranaire, en utilisant un large éventail de techniques, allant de la microscopie de fluorescence et de la RMN à l’état solide (ssNMR) à la microscopie de force atomique (AFM), la cryo-microscopie électronique (cryoEM) et la modélisation informatique. Nous souhaitons découvrir l’impact de l’oligomérisation et de la phosphorylation de StREM1.3 sur ses interactions membranaires et son activité biologique, ainsi que d’examiner son influence sur la dynamique des lipides et les lipides requis pour l’accrochage à la membrane et le regroupement en nanodomaines. Enfin, forts de toutes les données structurales disponibles, nous entreprendrons la reconstruction in vitro et la caractérisation de nanodomaines minimaux de StREM1.3Group 1 isoform 3 remorin from Solanum tuberosum (StREM1.3) is a membrane protein belonging to the multigenic family of plant proteins called remorins (REMs), involved in plant immunity, symbiosis, abiotic stress resistance and hormone signalling. REMs’ most well known feature is their ability to segregate into nanodomains at the plasma membrane’s (PM) inner leaflet. For StREM1.3, this is achieved by an interaction between two lysines of the remorin C-terminal anchor (RemCA) and negatively charged phosphatidylinositol 4-phosphate (PI4P). Thus, RemCA undergoes conformational changes and partially buries itself in the PM, resulting in an intrinsic membrane anchoring. Capitalising on pre-existing structural data about this isoform, we investigate StREM1.3’s membrane-interacting properties further, using a wide array of techniques, ranging from fluorescence microscopy and solid-state nuclear magnetic resonance (ssNMR) to atomic force microscopy (AFM), cryo-electron microscopy (cryoEM) and computational modelling. We aim to discover the impact of StREM1.3’s oligomerisation and phosphorylation on its membrane interactions and biological activity, and to assess its influence on lipid dynamics as well as its lipid requirements for membrane binding and nanoclustering. Finally, based on all available structural data, we will undertake the in vitro reconstruction and characterisation of minimal nanodomains of StREM1.3
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Yu, Chao. « Quantitative Study of Membrane Nano-organization by Single Nanoparticle Imaging ». Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLX054.
Texte intégralRésumé :
La nano-organisation de la membrane cellulaire est essentielle à la régulation de certaines fonctions cellulaires. Dans cette thèse, les récepteurs EGF, CPεT et de la transferrine ont été marqués avec des nanoparticules luminescentes et ont été suivis à la fois dans leur environnement local dans la membrane cellulaire vivantes pour de longues durées et sous un flux hydrodynamique. Nous avons alors appliqué des techniques d'inférence bayésienne, d’arbre de décision et de clustering de données extraire des informations quantitatives sur les paramètres caractéristiques du mouvement des récepteurs, notamment la forme de leur confinement dans des microdomaines. L’application d’une force hydrodynamique sur les nanoparticules nous a alors permis de sonder les interactions auxquelles ces récepteurs sont soumis. Nous avons appliqué cette approche in vitro pour favoriser et mesurer la dissociation in vitro de paires récepteur / ligand à haute affinité entre des récepteurs membranaires et leurs ligands pharmaceutiques, telles que HB-EGF et DTR et l’avons ensuite appliqué à l’étude d’interactions à la membrane cellulaire. Nous avons ainsi mis en évidence trois modes différents d'organisation de la membrane et de confinement des récepteurs: le confinement de CPεTR est déterminé par l'interaction entre les récepteurs et les constituants lipidiques / protéiques des microdomaines, le potentiel de confinement de l'EGFR résulte de l'interaction avec les lipides et les protéines de l’environnement du radeau et de l’interaction avec la F-actine; les récepteurs de la transferrine diffusent librement dans la membrane, uniquement limités stériquement par des barrières d’actine, selon le modèle ‘picket-and-fence’. Nous avons de plus montré que les nanodomaines de type radeau sont rattachés au cytoskelette d’actine. Ce travail présente donc à la fois un aperçu quantitatif du récepteur membranaire, des mécanismes d’organisation à l’échelle nanométrique, et établit un cadre méthodologique avec lequel différents types de propriétés membranaires peuvent être étudiésIn this thesis, EGF, CPεT and transferrin receptors were labeled with luminescent nanoparticles, , and were tracked both in their local environment in the cell membrane and under a hydrodynamic flow. Bayesian inference, Bayesian decision tree, and data clustering techniques can then be applied to obtain quantitative information on the receptor motion parameters. Furthermore, we introduced hydrodynamic force application in vitro to study biomolecule dissociation between membrane receptors and their pharmaceutical ligands in high affinity receptor- ligand pairs, such as HB-EGF and DTR. Finally, three different modes of membrane organization and receptor confinement were revealed: the confinement of CPεTR is determined by the interaction between the receptors and the lipid/protein constituents of the raft; the confining potential of EGFR results from the interaction with lipids and proteins of the raft environment and from the interaction with F-actin; transferrin receptors diffuse freely in the membrane, only sterically limited by actin barriers, according to the “picket-and-fence” model. We moreover showed that all raft nanodomains are attached to the actin cytoskeleton
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Hebisch, Elke [Verfasser], et Stefan W. [Akademischer Betreuer] Hell. « STED microscopy of cardiac membrane nanodomains / Elke Hebisch ; Betreuer : Stefan W. Hell ». Heidelberg : Universitätsbibliothek Heidelberg, 2017. http://d-nb.info/1180740068/34.
Texte intégral
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Hebisch, Elke [Verfasser], et Stefan [Akademischer Betreuer] Hell. « STED microscopy of cardiac membrane nanodomains / Elke Hebisch ; Betreuer : Stefan W. Hell ». Heidelberg : Universitätsbibliothek Heidelberg, 2017. http://nbn-resolving.de/urn:nbn:de:bsz:16-heidok-227475.
Texte intégral
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Liu, Xian-He. « Perfluoroalkylated compounds at the Interfaces : surface nanodomains and spherulites : interactions with phospholipid films ». Thesis, Strasbourg, 2018. http://www.theses.fr/2018STRAF029.
Texte intégralRésumé :
Cette thèse concerne l’auto-assemblage d'alcanes semi-fluorés (FnHm) et de fluorocarbures (FCs) aux interfaces et leurs interactions avec des phospholipides (PLs) en 2D et 3D. Nous avons étudié les sphérulites formées dans des films de diblocs FnHm. La morphologie de ces sphérulites, concentrique ou radiale, est contrôlée par la longueur des blocs Fn et Hm et la vitesse de refroidissement. Les nanodomaines de diblocs FnHm dans les monocouches de Langmuir sont incompressibles et forment des gels physiques 2D, même à pression nulle. Les blocs Hm sont cristallisés et inclinés d’ ~30°C par rapport à la normale à la surface. La réflectivité de neutrons a montré que l’albumine adsorbée sur des monocouches de PL est désorbée par un FC gazeux. Ce résultat pourrait permettre de lutter contre l’inactivation du surfactant pulmonaire par les protéines sériques. L’ajout de diblocs à des films de PLs accroit l'élasticité des monocouches. Nous avons préparé des microbulles stables à parois de PLs/FnHm, les diblocs agissant en co-surfactantsThis thesis focuses on the self-assembly of semi-fluorinated alkanes (FnHm) and fluorocarbons (FCs) at interfaces and their interactions with phospholipids (PLs) in 2D and 3D. 2D Spherulites were identified in FnHm films for the first time. Their morphology, ring-banded or radial, was controlled by varying block lengths and cooling rate. Nanodomains of FnHm in monolayers formed incompressible 2D physical gels, even at zero surface pressure. The Hm segments are crystalline and titled by 30°C to the normal to the surface. Neutron reflectivity showed that albumin adsorbed on PLs monolayers is desorbed by exposure to FC gas, which opens the potential use of FCs to treat the inactivation of the lung surfactant by serum proteins. Incorporating FnHm into PL monolayers increases their elasticity. Small, stable microbubbles of PLs/FnHm were obtained. FnHm diblocks function as co-surfactants for stabilizing microbubbles
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Parutto, Pierre. « Statistical analysis of single particle trajectories reveals sub-cellular nanodomain organisation and function ». Thesis, Paris Sciences et Lettres (ComUE), 2019. http://www.theses.fr/2019PSLEE055.
Texte intégralRésumé :
Les trajectoires de molécules individuelles obtenues par microscopie super-résolution permettent de suivre des protéines avec une précision nanométrique dans des cellules vivantes. Dans cette thèse, j’ai étudié les régions de hautes densités présentes dans ces trajectoires, dont un modèle possible est celui des puits de potentiel. Pour les caractériser à partir de trajectoires, j’ai développé une nouvelle méthode hybride basée sur la densité de points et le champ de force local puis je l’ai comparé aux méthodes d’état de l’art. Ensuite, j’ai utilisé celle-ci pour caractériser les puits maintenant les canaux calciques Cav au niveau des zones actives des terminaux présynaptiques ce qui a permis de mieux comprendre le rôle des variantes d’épissage de ces canaux dans la transmission synaptique. Dans une autre étude, j’ai analysé des trajectoires de protéines résidant dans le lumen du Réticulum Endoplasmique (RE). J’ai créé une méthode pour reconstruire le réseau du RE à partir des trajectoires que j’ai utilisé pour caractériser le mouvement de ces molécules par un modèle de saut-diffusion qui a pour conséquence une meilleure redistribution du contenu luminal par rapport à un mouvement diffusif. Enfin, je discute d’autres analyses de trajectoires pour les intéractions lysosome-ER, les canaux Cav à la jonction neuro-musculaire de la drosophile et les protéines composant le complexe NuRDSingle-Particle Trajectories (SPTs) obtained from super-resolution microscopy allow to track proteins with nanometer precision in living cells and are used in neuroscience and cellular biology. In this thesis, I was interested in the high-density nanodomains found in these trajectories that can be modeled as potential wells. To characterize them, I developed a new hybrid method based on the point density and local drift field and compared it to the other state-of-the-art methods. Then, I used it to identify transient potential wells in SPTs of voltage-gated calcium channels (CaV) contributing to a better understanding of the role of the different CaV splice variants in synaptic transmission. In another study, I looked at SPTs from Endoplasmic Reticulum (ER) luminal resident proteins where I developed a method to reconstruct the network from trajectories and used it to characterize the luminal motion as a jump-diffusion process, which allows for a better redistribution of the luminal content than the previously assumed diffusive model. Finally, I discuss other analyses of motions for lysosome-ER interactions, CaV2.1 channels at drosophila’s neuromuscular junctions and the description of the motion of the constituent proteins of the NuRD chromatin remodeling complex
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Sachl, Radek. « Localisation of Fluorescent Probes and the estimation of Lipid Nanodomain sizes by modern fluorescence techniques ». Doctoral thesis, Umeå universitet, Kemiska institutionen, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-52619.
Texte intégralRésumé :
The thesis is divided into two major parts. The first part focuses on the localisation of probes in lipid/polymeric bilayers and in GM1 micelles. Included in this thesis is a new approach based on electronic energy transfer/migration (FRET/DDEM), which efficiently determines transversal positions of fluorescent molecules in lipid bilayers. This approach has been used to locate newly synthesized lipid probes in DOPC bilayers. The label was introduced at the end of sn-2 acyl chains of variable length. Analytical models accounting for FRET exist for a limited number of basic geometries. Here, a combination of FRET and Monte Carlo simulations enables the localisation of probes in bicelles and in bilayers containing pores, i.e. in lipid systems with variable curvature, or in non-homogenous lipid systems. This approach has been used to test whether conical-like fluorescence probes have an increased affinity to highly curved regions, which would enable preferential labelling of membrane pores. A simplified FRET model has been applied to localize 2-pyridones, a class of potential drugs, in GM1 micelles. Since the localisation of drugs within nanoparticles might influence the release kinetics and loading efficiency, knowledge about the drug location is highly relevant. It turned out that all derivatives were localised at the core-shell interface of GM1 micelles. The second part of the thesis focuses mainly on the estimation of lipid nanodomain size by means of FRET, which still remains the most powerful method in this field. Limitations of FRET in the determination of domain size have been explored. We showed that the limitations of FRET are mainly caused by a low probes affinity to either the liquid-ordered or liquid-disordered phase. In the continuing work we provided a detailed dynamic and structural study of crosslinker-triggered formation of nanodomains. Here, two different domains have been revealed, i.e. i) domains whose size grows with increasing amount of added cholera toxin (CTxB), and to which CTxB binds tightly; ii) domains formed in membranes containing a slightly increased amount of sphingomyelin (as compared to i) whose size does not change during titration by additional CTxB and to which CTxB binds less tightly.Disertace je rozdělena do dvou hlavníchčástí. Prvníčást se zabývá lokalizací značek v lipidových/polymerních dvojvrstvách a v GM1micelách. V práci prezentujeme nový přístup založený na přenosu/migraci elektronické energie (FRET/DDEM), jež umožňuje efektivně určovat vertikální pozici fluorescenčních molekul uvnitř lipidové dvojvrstvy. Tato metoda byla použita k lokalizaci nově syntetizovaných lipidových značek značených na konci sn-2 acylového řetězce s různou délkou v DOPC dvojvrstvách. Analytické modely popisující FRET existují pouze pro limitovaný počet základních geometrií. Kombinace FRETu s Monte Carlo simulacemi nicméně umožňuje lokalizaci značek v bicelách a v dvojvrstvách obsahujících póry, tj. v lipidových systémech s proměnlivým zakřivením a v nehomogenních lipidových útvarech. Tento přístup umožnil např. zjistit, zda kuželovitětvarované značky mají zvýšenou afinitu k vysoce zakřiveným oblastem dvojvrstvy, což by umožnilo preferenční značení pórů. Lokalizovány byly rovněž tři deriváty 2-pyridonů(potencionálních léčiv) v GM1micelách za použití jednoduchého modelu zohledňujícího FRET mezi donory a akceptory nacházejícími se v micelách. Lokalizace léčiv v nanočásticích ovlivňuje kinetiku uvolňování (release kinetics) a množství látky solubilizované v micelách (loading efficiency). Druhá část se především zabývá určováním velikostí lipidových nanodomén pomocí FRETu, který stále zůstává nejvíce výkonnou metodou v této oblasti. Zkoumány byly limitace FRETu v určování lipidových nanodomén. Ukázalo se, že tato omezení jsou především způsobena nízkou afinitou značek buď k Lonebo k Ldfázi. V navazující studii jsme poskytnuli detailní dynamickou a strukturní studii formace nanodomén indukované crosslinkerem. Objevili jsme dva typy domén: a) domény, jejichž velikost se zvětšuje s rostoucím množstvím přidaného cholera toxinu (CTxB) a k nimž se CTxB váže pevně a b) domény vzniklé v membránách se zvýšeným množstvím sfingomyelinu (ve srovnání s a)), jejichž velikost se nemění během titrace dodatečným CTxB a k nimž se CTxB váže méně pevně.
This thesis has been elaborated within the framework of the Agreement on JointSupervision (co-tutelle) of an International Doctoral Degree Programmebetween Charles University in Prague, Czech Republic and the Department of Chemistry at Umeå University, Sweden.
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Kirsch, Sonja [Verfasser], Rainer [Akademischer Betreuer] Böckmann et Rainer [Gutachter] Böckmann. « The Role of Membrane Nanodomains in Permeation / Sonja Kirsch ; Gutachter : Rainer Böckmann ; Betreuer : Rainer Böckmann ». Erlangen : Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), 2019. http://d-nb.info/1196875901/34.
Texte intégral
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Yandrapalli, Naresh. « Role of HIV-1 Gag protein multimerization in the generation of nanodomains in lipid membranes ». Thesis, Montpellier, 2016. http://www.theses.fr/2016MONTT097/document.
Texte intégralRésumé :
La polyprotéine Gag du VIH-1 qui contient quatre principaux domaines (Matrix (MA), capside (CA), nucléocapside (NC), et P6) est l’orchestrateur privilégié de l'assemblage du virus HIV-1, assemblage qui a lieu pendant la phase tardive de la réplication. Il est bien connu que Gag interagit avec les lipides de la membrane plasmique de la cellule hôte et s’auto-assemble sur le feuillet interne de cette dernière afin de générer de nouvelles particules virales. Le bourgeonnement de ces particules virales hors de la cellule hôte est décrit comme étant dépendant de la machinerie cellulaire ESCRT. Différentes études structurales, fonctionnelles ainsi que des simulations de dynamique gros grain ont montré que la liaison de Gag à la membrane est médiée par une interaction duale. Une spécifique de nature éléctrostatique, qui associe une région hautement basique (HBR) du domaine MA de Gag au lipide acide,phosphatidyl inositol biphosphate (PI(4,5)P2) du feuillet interne de la membrane plasmique. Une de type hydrophobe, qui consiste en l’insertion du myristate de Gag dans la membrane plasmique. Savoir si Gag reconnait spécifiquement des domaines lipidiques pré-existants de type « rafts » ou si, au contraire, Gag tri ses lipides et les réorganise latéralement afin d’optimiser sa multimérisation et son bourgeonnement est une question à la fois fondamentale et d’actualité en virologie.Durant ma thèse, j’ai vérifié l’existence de la seconde hypothèse en utilisant des membranes modèles contenant du PI (4,5) P2 marqué de façon fluorescente et différent mutants et produits de la protéine Gag non-myristoylée. Ces expériences ont montré de fortes affinités de ces protéines pour les membranes contenant du PI (4,5) P2. S’appuyant sur les propriétés d’auto-extinction de fluorescence du marqueur choisit et à l’aide des différents variants de la protéine Gag, j'ai pu montré que la multimérisation de Gag génère l’existence de nanodomaines contenant du PI (4, 5) P2 et du cholestérol, la sphingomyéline étant au contraire exclue de ces domaines. En marquant la protéine Gag par un autre fluorophore, j’ai pu montrer par microscopie optique sur des vésicules lipidiques géantes (GUVs) que la protéine Gag partitionnait préférablement dans des microdomaines lipidiques de type liquide désordonnés (Ld). Par la suite, j’ai testé la capacité de la protéine Gag d’induire la formation de vésicules sur des membranes modèles (Bicouches supportés et GUVs) contenant du PI(4,5) P2 et de la phosphatidyl sérine (PS). En utilisant une microbalance à cristal de quartz (QCM-D) et des techniques de microscopie de fluorescence, j’ai suivi l'auto-assemblage de Gag dans le temps et ai montré que la protéine Gag était suffisante pour générer une courbure de la membrane et libérer des vésicules lipidiques. Grâce à différents produits de maturation de cette protéine, j’ai montré que la présence des domaines MA et CA est suffisante pour produire ces vésicules.L’ensemble de ces résultats suggèrent que la liaison et la multimérisation de la protéine Gag ne se produit pas dans des domaines lipidiques préexistants de type « raft », mais, au contraire, que la liaison et multimérisation de la protéine Gag génère l’existence de domaines lipidiques enrichis en PI (4,5) P2 et en cholestérol. La générescence de ces domaines lipidiques pourrait participer à la courbure de la membrane plasmique nécessaire au bourgeonnement du virusGag polyprotein of HIV-1 is made of four main domains Matrix (MA), Capsid (CA), Nucleocapsid (NC), and P6 and is the prime orchestrator of virus assembly that occurs during the late phase of replication. It is well known that Gag interacts with host cell lipids and self-assemble along the inner-leaflet of the plasma membrane in order to generate virus like particles (VLPs). Budding of these VLPs out of the living cell is described to be ESCRT dependent. Structural, functional and simulation based studies has shown that Gag membrane binding is mediated by a bipartite interaction. One specific electrostatic interaction, between the highly basic region (HBR) of its MA domain and the host cell acidic lipid phosphatidyl inositol bisphophate (PI(4,5)P2), plus a hydrophobic interaction through Gag’s myristate insertion in the plasma membrane. It is still an opened question whether Gag would specifically recognize pre-existing lipid domains such as rafts to optimize its multimerization or, on the contrary, would reorganize lipids during its multimerization. During my Ph.D. I explored the second hypothesis using purified myr(-) Gag protein and model membranes containing fluorescently labelled PI(4,5)P2.Bonding experiments have shown strong affinities of these purified proteins towards PI(4,5)P2 containing lipid bilayers. Using PI(4,5)P2 fluorescence self-quenching properties, I found that multimerization Gag generates PI(4,5)P2/Cholesterol enriched nanoclusters. On the opposite, sphingomyelin was excluded from these nanoclusters. In addition to this, using a fluorescently labelled myr(-) Gag, I have observed its preferable partitioning into lipid disordered (Ld) phases of giant unilamellar vesicles (GUVs). Further, possibility of whether HIV-1 Gag alone, as a minimal system, can induce the formation of vesicles on PI(4,5)P2/PS containing supported lipid bilayers (SLBs) & GUVs was tested. Using quartz crystal microbalance (QCM-D) and fluorescence microscopy techniques, I monitored the self-assembly of HIV-1 Gag with time and found that Gag was sufficient to generate membrane curvature and vesicle release. Moreover, using mutants of this protein, I found that having MA and CA domain is enough for Gag to produce vesicle like structures. Taken together, these results suggest that binding and multimerization of Gag protein does not occur in pre-existing lipid domains (such as “rafts”) but this multimerization is more likely to induce PI(4,5)P2/Cholesterol nanoclusters. This nanophase separation could locally play a role in the membrane curvature needed for the budding of the virus
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Sota, Norihiro. « Elucidation of Self-Assembling Mechanism and Dynamics in Block Copolymer Nanodomain Structures Induced by Phase Transitions ». 京都大学 (Kyoto University), 2013. http://hdl.handle.net/2433/174981.
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Zandt, Maximilian [Verfasser]. « Nanodomain clustering mechanisms of Junctophilin-2 in human kidney, cardiac and skeletal muscle cells / Maximilian Zandt ». Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2020. http://d-nb.info/1220080632/34.
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Kirsch, Wolfgang. « Biophysical analysis of the spatial and temporal distribution of free Ca2+ ions within micro- and nanodomains of muscle cells ». [S.l.] : [s.n.], 2000. http://deposit.ddb.de/cgi-bin/dokserv?idn=961763817.
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Schlegel, Jan [Verfasser], Markus [Gutachter] Sauer, Ulrich [Gutachter] Terpitz, Christian [Gutachter] Wegener et Katrin [Gutachter] Heinze. « Super-Resolution Microscopy of Sphingolipids and Protein Nanodomains / Jan Schlegel ; Gutachter : Markus Sauer, Ulrich Terpitz, Christian Wegener, Katrin Heinze ». Würzburg : Universität Würzburg, 2021. http://d-nb.info/1229352414/34.
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Liang, Pengbo [Verfasser], et Thomas [Akademischer Betreuer] Ott. « The role of membrane nanodomains and the cell wall-plasma membrane-cytoskeleton continuum during symbiotic infection in Medicago truncatula ». Freiburg : Universität, 2020. http://d-nb.info/1220631760/34.
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Gao, Yan [Verfasser], Ralf [Akademischer Betreuer] Riedel et Wolfgang [Akademischer Betreuer] Ensinger. « Nanodomain Structure and Energetics of Carbon Rich SiCN and SiBCN Polymer-Derived Ceramics / Yan Gao. Betreuer : Ralf Riedel ; Wolfgang Ensinger ». Darmstadt : Universitäts- und Landesbibliothek Darmstadt, 2014. http://d-nb.info/1108094201/34.
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Kirsch, Wolfgang G. « Biophysical analysis of the spatial and temporal distribution of free Ca 2+ ions within micro- and nanodomains of muscle cells ». [S.l. : s.n.], 2001. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB8986367.
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Guyomard, Aurélie. « Elaboration de films polyelectrolytes a base de polysaccharides amphiphiles : application à l’immobilisation de composés hydrophobes d’intérêt biologique ». Rouen, 2007. http://www.theses.fr/2007ROUES064.
Texte intégralRésumé :
Dans le but d’immobiliser des composés hydrophobes présentant un intérêt biologique à l’interface solide/liquuide, des fims minces à base de polysaccharides amphiphiles et de divers polycations ont été élaborés par la technique d’auto-assemblage couche par couche (lbl). Cette technique consiste à adsorber successivement sur un support chargé des couches de polycations et de polyanions. Les polysaccharides utilisés sont des carboxyméthylpululanes (CMP) porteurs de chaînons alkyle (CMP-τCn). L’influence des caractéristiques moléculaires eet macromoléculaires de ces dérivés (leur hydrophobie, leur masse molaire, etc) sur les caractéristiques des films réalisés a été systématiquement étudiée. Il a été notamment montré qu’il était possible de moduler l’incrément d’épaisseur ainsi que la microstructure des films polyélectrolytes en jouant sur le caractère plus ou moins hydrophobe des polysaccharides amphiphiles. Des corrélations ont été établies entre les propriétés des polysaccharides amphiphiles en solution et les caractéristiques des films obtenus. Il a été montré que les nanodomaines hydrophobes intra et/ou intermoléculaires formés en solution aqueuse étaient préservés lors du processus d’auto-assemblage. Ces nanodomaines ont été utilisés comme nano-réservoirs pour le piégeage de colorants hydrophobes. Il a été montré que la capacité de piégeage des films pouvait être modulée en fonction du caractère plus ou moins hydrophobes des carboxyméthylpullulanes alkylés. Enfin, un peptide hydrophobe ionophore, la gramicidine A, a été incorporé avec succès dans ces assemblages polyélectrolytes dans le but de préparer des films antibactériens. L’activité de ces derniers a été vérifiée auprès d’une bactérie Gram+, Enterococcus FaecalisThe goal of the project was to immobilize hydrophobic drugs or "biomolecules" in thin films containing amphiphilic polysaccharides. The films were obtained using the Layer-by-layer technique (LbL). This technique consists of sequential adsorption of polycations and polyanions onto a charged substrate. The modified polysaccharides used in this study are obtained by grafting alkyl chains on carboxyméthylpullulan (CMP) precursor. The influence of the polyelectrolytes characteristics used on the films growth was systematically studied. We showed that it was possible to modulate the thickness as well as the multilayers films microstructure by changing the characteristics of the amphiphilic polysaccharides used. Moreover the properties of amphiphilic polysaccharides in solution and at solid/liquid interfaces are similar. Furthermore the intra and/or intermolecular hydrophobic nanodomains are preserved during adsorption process. These hydrophobic nanodomains showed their potential as nanocontainers for actives molecules or dyes. Moreover the sequestration efficiency of the films can be modulated as a function of the hydrophobicity of the alkylated CMP. Lastly, an hydrophobic ionophore antibacterial polypeptide "gramicidin A" was trapped in these nanocontainers and its antibacterial activity against Enterococcus faecalis was tested. Our results clearly show that this technique of incorporation of an active species within multilayers film allow us to obtain an antibacterial surface by a simple way
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Mohamedgamil, Sana Siddig Abdelrahman [Verfasser], et Davide [Gutachter] Calebiro. « Organization and dynamics of class C GPCR nanodomains in neurons visualized by single-molecule microscopy / Sana Siddig Abdelrahman Mohamedgamil ; Gutachter : Davide Calebiro ». Würzburg : Universität Würzburg, 2020. http://d-nb.info/1207760897/34.
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Deroubaix, Anne-Flore. « Rôle de la rémorine et des nanodomaines membranaires dans la signalisation de la réponse aux phytovirus ». Thesis, Bordeaux, 2019. http://www.theses.fr/2019BORD0292.
Texte intégralRésumé :
Dans la lutte contre les virus, les plantes ont mis au point divers mécanismes de défenses pour se protéger contre les agents pathogènes. Les protéines végétales localisées à la membrane plasmique telles que les rémorines (REM) peuvent limiter l’infection virale. Les REM appartiennent à une famille de protéines multigéniques spécifiques à la plante, classées en six groupes phylogénétiques localisées dans les nanodomaines de la membrane plasmique et, dans certains cas, au niveau des plasmodesmes. Notre équipe avait précédemment montré que chez la tomate et Nicotiana benthamiana, la surexpression de l’isoforme 3 du groupe 1 de Solanum tuberosum (StREM1.3) limitait la propagation de cellule-à-cellule du Potato Virus X, un Potexvirus sans affecter la réplication virale. Au cours de ma thèse, nos données ont permis de construire un modèle de travail dans lequel une protéine kinase dépendante du calcium (AtCPK3) d’Arabidopsis thaliana est capable d’interagir in vivo avec StREM1.3, de phosphoryler le domaine N-terminal de StREM1.3 et, enfin, avec l'aide de protéines non caractérisées à ce jour, conduire à la restriction du mouvement de cellule à cellule de PVX chez N.benthamiana. N.benthamiana , parfaite pour l'expérimentation virale, est allo-tétraploïde, rendant de ce fait difficiles les études génétiques. Sachant qu’il existe 34 isoformes de CPKs, avec une redondance fonctionnelle probable entre elles, nous avons basculé sur un autre pathosystème : nous avons choisi Arabidopsis thaliana profitant de la « boîte à outils » génétiques que cette plante offre et nous avons choisi une autre espèce de virus, de la famille des potexvirus capable d'infecter A. thaliana : le Plantago Asiatica Mosaic Virus (PlAMV). Les objectifs sont 1 / d’étudier la contribution des différents clades de REM dans le mouvement intercellulaire des potexvirus 2 / comprendre quels CPK sont impliquées dans ce processus en utilisant les mutants simples et double de REM mais aussi de CPK, ainsi que les surexpresseurs AtCPK 3 / Etudier la contribution du groupe 1 de REM et de CPK3 dans le mouvement systémique du potexvirus. Nous avions précédemment montré que, comme le PVX, le mouvement local du PlAMV est limité par StREM1.3 et AtCPK3 dans N.benthamiana. Nous avons optimisé les conditions expérimentales pour suivre et comparer le PlAMV marqué GFP dans différents fonds génétiques d'Arabidopsis. En utilisant cette méthode, nous avons pu suivre à la fois le mouvement de cellule à cellule du virus localement, puis l’infection systémique à travers la plante entière. Les mutants knock-out simples et multiples du groupe 1 de REM, ainsi que les surexpresseurs CPK ont été utilisés. Fait intéressant, nous n'avons pas détecté de différence de propagation par rapport au contrôle sur divers mutants de CPK, sauf dans le mutant cpk3KO. En effet, tant au niveau local que systémique, la propagation du PlAMV est améliorée sur le mutant cpk3KO alors que les lignées surexprimant CPK3 présentent un effet opposé, démontrant la grande implication de CPK3 dans la propagation du potexvirus. De même, nous démontrons la redondance de chaque isoforme du groupe 1 de REM sur la restriction du mouvement intercellulaire du PlAMV. Il est intéressant de noter que REM favorise la propagation intercellulaire d’un autre genre viral, le genre Potyvirus, ce qui suggère que les fonctions de REM ne sont pas généralisables pour tous les genres. Globalement, nos résultats classifient les groupes 1 de REM et CPK3 en tant que protéines de défenses antivirales dans les infections aux potexvirus, que ce soit au niveau local ou systémique, et suggèrent que la fonction de REM est dépendante du genre viral. Cette recherche ouvrira la voie sur de nouvelles clés thérapeutiques, pour in fine, lutter contre les infections viralesIn the battle against viruses, plants have evolved various defence mechanisms to protect themselves against pathogens. Membrane-bound plant proteins such as Remorin (REM) may restrict viral infection. REMs belong to a plant-specific multigene family, classified in six phylogenetic groups that are localized in plasma membrane nanodomains and for some of them in plasmodesmata. Our team previously showed that in tomato and Nicotiana benthamiana, overexpression of Solanum tuberosum group 1 isoform 3 (StREM1.3) limits the cell-to-cell spread of the potexvirus Potato virus X (PVX) without affecting viral replication. During my thesis, our data allowed to built a working model in which the Arabidopsis thaliana CALCIUM-DEPENDENT PROTEIN KINASE 3 (AtCPK3) is able to interact with group 1 REM in vivo, phosphorylates the N-terminal domain of StREM1.3 and, finally, with the help of uncharacterized proteins lead to the restriction of PVX cell-to-cell movement in N.benthamiana. N.benthamiana is perfect for viral experimentation, but is allo-tetraploid, making it difficult for genetic studies. Because of CPKs have 34 isoforms with likely functional redundancy between them, we switched to another pathosystem using the genetic toolbox of Arabidopsis thaliana and a potexvirus species able to infect A. thaliana, the Plantago Asiatica Mosaic Virus (PlAMV). The objectives are 1/ to study the contribution of different REM clades in potexvirus intercellular movement; 2/ to understand which CPKs are involved in this process using REM and CPKs single and multiple mutants, as well as AtCPKs over-expressors; 3/ To study the contribution of Group 1 REM and CPK3 on systemic potexvirus movement. We previously showed that, like PVX, PlAMV local movement is restricted by StREM1.3 and AtCPK3 in N.benthamiana. We optimized the experimental conditions to track and compare GFP-tagged PlAMV in different Arabidopsis genetic backgrounds. By using this method, we were able to track both local virus cell-to-cell movement and systemic infection through the whole plant. Group 1 REM and CPK single and multiple knock out mutants, as well as CPK over-expressors wereused. Interestingly, we did not detect any difference in propagation compared with control on various CPKs KO, except in cpk3 mutant. Indeed, both in local and systemic, PlAMV propagation is enhanced on cpk3 mutant while CPK3 overexpressing lines display an opposite effect, demonstrating the great involvement of CPK3 in potexvirus propagation. Similarly, we demonstrate the redundancy of each isoform from group 1 REM on the restriction of the intercellular movement of PlAMV. Interestingly, REM promotes intercellular propagation of another viral genus, the potyvirus genus, suggesting that REM functions are not general for all genera. Globally, our results classify group 1 REM and CPK3 as antiviral defence protein both in local and systemic potexvirus infection, and suggest that REM function is viral genus dependent. This research will pave the way toward new host targets to fight phytovirus infection
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Kofahl, Claudia. « Oberflächenstrukturen modulierter Systeme - Darstellung von regelmäßig angeordneten, polaren Nanodomänen mittels Piezoresponse Force Microscopy ». Doctoral thesis, Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2020. http://hdl.handle.net/21.11130/00-1735-0000-0005-1430-E.
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Peng, Jiangguli. « Effect of the microstructure and orientation of grains on the performance of perovskite ferroelectric ceramics ». Thesis, Le Mans, 2020. http://www.theses.fr/2020LEMA1013.
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Les matériaux ferroélectriques sont largement utilisés dans d’importantes technologies incluant les transducteurs, actionneurs, capteurs... Parmi les structures ferroélectriques les plus exploitées, celles à base de Pb(Zr,Ti)O3 (PZT) offrent de bonnes performances dans des dispositifs opérationnels. Malgré cet aboutissement en terme de valorisation, la compréhension fondamentale des caractéristiques physiques des systèmes basés sur les céramiques PZT continue de susciter l’intérêt d’une large communauté scientifique. Cependant, un intérêt croissant porte sur la mise en œuvre de structures ferroélectriques exemptes d’éléments chimiques nocifs pour l’environnement dont la teneur en plomb. Dans ce cadre, les systèmes à base de BiFeO3-BaTiO3 (BF-BT) ont suscité beaucoup d'intérêt en tant que matériaux ferroélectriques sans plomb. Ce travail de thèse porte sur des études systématiques portant sur les céramiques à base de PZT et celles à base de BF-BT. Trois contributions ont été développées doant la première dédiée à l'analyse des microstructures, des phases cristallines et des propriétés ferroélectriques dans les systèmes PZT dopés. Les travaux ont montré le rôle du dopage Soft-Hard sur les propriétés structurales, morphologiques et électriques des matériaux dopés accepteurs. Dans une deuxième partie, l'interaction des microstructures et des propriétés électriques a été étudiée dans les systèmes BF-BT. L’optimisation des conditions de fabrication à travers la composition chimique et le traitement thermique contribue aux propriétés piézoélectriques améliorées dans ces systèmes sans plomb. Pour la troisième partie des travaux, des études expérimentales de fabrication, texturation et les caractérisations des propriétés ferroélectriques et électromécaniques ont été corrélées avec l’organisation structurale et morphologique des céramiques texturéesFerroelectric materials have been widely applied in transducers, high-pressure generators, actuators, sensors... In this context, the ferroelectric materials are generally regarded as being related to an important class of smart materials. Among the most popular ferroelectric materials, those based on Pb(Zr,Ti)O3 (PZT) structures show the best performances in operating devices. Despite such achievements, the fundamental understanding of the physical characteristics continues to arouse the interest of a wide scientific community. On the other hand, in view of the environmental-friendly requirement, more attention have been paid by the researchers to lead-free ferroelectric materials. Thus, BiFeO3-BaTiO3 (BF-BT) systems have attracted a great deal of interest as promising candidate for lead-free ferroelectric materials. In this dissertation, PZT-based and BF-BT-based ceramics were investigated systematically. Three contributions have been developed with the first dealing with the analysis of microstructures, crystalline phases and electric properties in doped PZT systems. It was revealed the involvement of softening-hardening features and the relation between defect dipoles and electric properties in acceptor doped materials. In a second part, the interplay of microstructures and electric properties was studied in BF-BT systems, providing the relevant analysis of the optimized performance such as the enhanced piezoelectric properties of lead-free systems. In the third contribution, experimental methods of synthesis and texturing BF-BT systems by BT templates ad the characterization of ferroelectric and electromechanical properties have been correlated with the structural and morphological organization of textured ceramics
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Noack, Lise. « Rôle du complexe AtPI4Kalpha1 dans l’établissement de l’identité de la membrane plasmique et le développement chez Arabidopsis thaliana ». Thesis, Lyon, 2020. http://www.theses.fr/2020LYSEN066.
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Les cellules sont composées de compartiments délimités par une membrane. Pour permettre aux protéines d’être associées aux membranes du bon compartiment, chaque membrane a une identité qui lui ait propre. Elle se définit par ses caractéristiques physiques et chimiques. Cependant, les compartiments d’une cellule échangent du matériel en permanence. Comment l’identité des membranes est maintenue malgré le flux constant d’échanges de protéines et de lipides lors des transports vésiculaires et non-vésiculaires ? Les phosphoinositides (PIPs) sont des lipides anioniques présents en faible quantité dans les membranes. Chaque PIP se localise différemment dans la cellule. Parmis les PIPs, le PI4P est présent à la membrane plasmique et au Golgi/trans-Golgi Network (TGN). Chez les plantes, PI4P est majoritairement trouvé à la membrane plasmique, contrairement aux animaux ou à la levure chez qui le PI4P se trouve principalement au niveau du TGN. Ainsi le gradient de PI4P le long de la voie d’endocytose est inversé chez les plantes par rapport aux autres eucaryotes. Chez les plantes, l’accumulation de PI4P à la membrane plasmique confère un champ électrostatique important qui recrute des protéines spécifiquement à cette membrane. Comment le gradient de PI4P est établi chez les plantes ? PI4Kα1 est capable de produire du PI4P à partir de phosphatidylinositol. PI4Kα1 se localise à la MP. Par ailleurs, PI4Kα1 est la sous-unité catalytique d’un complexe comprenant 3 autres protéines : NO POLLEN GERMINATION (NPG), EFR3 OF PLANTS (EFOP) et HYCCIN. Le complexe est ciblé à la membrane plasmique via une ancre lipidique au niveau de EFOP. Les orthologues de PI4Kα1 chez la levure et les animaux sont localisés à la membrane plasmique par des complexes protéiques similaires, démontrant une conservation des mécanismes de production des PIP chez l’ensemble des eucaryotes. L’absence du complexe PI4Kα1 entraine une létalité de la plantes au niveau du grain de pollen ou de l’embryon. Ainsi PI4Kα1 est essentiel à l’identité de la membrane plasmique et par conséquent au développement de la planteEukaryotic cells are composed of several membrane-surrounded compartments. Each compartment has a unique physicochemical environment delimited by a membrane with a specific biochemical and biophysical identity. The membrane identity includes the nature of the lipids, the curvature, the electrostaticity and the density of lipids at the membrane. The identity of each membrane allows the proper localization of membrane-associated proteins. Phosphoinositides are rare anionic lipids present in membranes. Five types of phosphoinositides exist in plants - PI3P, PI4P, PI5P, PI(4,5)P2 and PI(3,5)P2 - depending of the number and the position of phosphates around the inositol ring. They accumulate differently at the plasma membrane and in intracellular compartments and interact with proteins through stereo-specific or electrostatic interactions. Recent work uncovered that PI4P concentrates according to an inverted gradient by comparison to their yeast and animal counterpart. In plants, PI4P massively accumulates at the plasma membrane and is present in fewer amounts at the trans-Golgi Network (TGN). This PI4P accumulation at the cell surface drives the plasma membrane electrostatic field, which in turn recruits a host of signalling proteins to this compartment. Moreover the plant TGN is the place of vesicular secretion but is also involved in endocytic sorting and recycling, which might imply regulatory mechanisms of lipid exchanges or membrane identity maintenance between the plasma membrane and the TGN. Here, we characterized PI4Kα1 mutants and showed that pi4kα1 loss-of-function leads to pollen grain lethality and distortion in the allele transmission via the female gametophyte, while its knockdown displayed strong developmental phenotypes. Using yeast two hybrid screening and mass spectrometry, we identified that PI4Kα1 is part of an heterotetrameric complex composed of NO POLLEN GERMINATION (NPG), EFR3 OF PLANTS (EFOP) and HYCCIN (HYC). The interaction between PI4Kα1 and the structural subunits of the complex is essential to target PI4Kα1 at the plasma membrane. In addition, we showed that PI4Kα1 complex is anchored in immobile and predefined subdomains of the plasma membrane. This work opens new perspectives on the role of the PI4Kα1 complex in plasma membrane suborganization
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Carsí, Rosique Marta. « Molecular mobility. Structure-property relationship of polymeric materials ». Doctoral thesis, Universitat Politècnica de València, 2016. http://hdl.handle.net/10251/59460.
Texte intégralRésumé :
[EN] The present work examines the influence of the chemical structure of polymers on thermal, mechanical and dielectric behavior. The experimental techniques used for the purpose are differential scanning calorimetry, dynamo-mechanical analysis and dielectric spectroscopy. Additionally, in order to confirm the results obtained using the above methods, other techniques such as ray diffraction have also been employed.
Chapters 1 and 2 contain the introduction and the objectives, respectively. Chapter 3 briefly describes the experimental techniques used.
Chapter 4 contains the findings of the comparative analysis of the response to electrical noise fields for three poly(benzyl methacrylates) with different structures. The analysis was carried out under a wide range of frequencies and temperatures on three poly(benzyl methacrylates) containing two dimethoxy groups in positions 2,5-, 2,3- and 3,4-. The results show that the position of the dimethoxy groups on the aromatic ring has a significant effect on the molecular dynamics of poly(benzyl methacrylate). The spectra obtained were of high complexity and therefore, in order to perform a better analysis, numerical methods for time-frequency transformation including the use of parametric regularization techniques were used. We studied the effect of this structural change on the secondary relaxation processes and relaxation process , relating to the glass transition. We also analyzed the effect of the dimethoxy group position on the formation of nanodomains, in which the side chains are predominant, and on the conduction processes of the materials tested.
In Chapter 5, the conductivity of rubbery liquids was studied by analyzing poly(2,3-dimethoxybenzyl methacrylate), which exhibits its own particular behavior. The chapter analyzes the principle of time-temperature superposition, employing different interrelated variables.
Chapter 6 focuses on how the presence of crosslinking affects the molecular mobility of polymethacrylates containing aliphatic alcohol ether residues. In this case, the effect of crosslinking on the secondary and primary relaxation processes was analyzed. The creation of nanodomains in the side chains as a result of the presence of crosslinking was also studied.[ES] En este trabajo se presenta un estudio de la influencia de la estructura química de los polímeros en su comportamiento térmico, mecánico y dieléctrico. Las técnicas experimentales empleadas para ello han sido la calorimetría diferencial de barrido, el análisis dinamo-mecánico y la espectroscopia dieléctrica. Adicionalmente, se han empleado otras técnicas como la difracción de rayos, con objeto de corroborar los resultados obtenidos por las primeras. En los Capítulos 1 y 2 se recoge la introducción y los objetivos, respectivamente. El Capítulo 3 presenta una breve descripción de las técnicas experimentales empleadas. En el Capítulo 4 se recogen los resultados obtenidos en el análisis comparativo de la respuesta a campos de perturbación eléctrica en un amplio rango de frecuencias y temperaturas para tres polimetacrilatos de bencilo con dos grupos dimetoxi en posiciones 2,5-, 2,3- y 3,4-. Los resultados obtenidos señalan el importante efecto de la posición de los grupos dimetoxi en el anillo aromático, sobre la dinámica molecular del polimetacrilato de bencilo. Los espectros obtenidos fueron muy complejos, por ello en orden a llevar a cabo un mejor análisis se emplearon métodos numéricos para la transformación tiempo-frecuencia que incluyeron el uso de técnicas de regularización paramétrica. Se ha estudiado el efecto que dicho cambio estructural ejerce tanto sobre los procesos de relajación secundaria como sobre el proceso de relajación α, relacionado con la transición vítrea. Así mismo, se ha analizado el efecto de la posición de los grupos dimetoxi en la formación de iii nanodominios en los que predominan las cadenas laterales, y su efecto en los procesos de conducción de los materiales analizados. En el Capítulo 5 se recoge el estudio de la conductividad de líquidos gomosos tomando como modelo el poli (metacrilato de 2,3-dimetoxibencilo), por su peculiar comportamiento. En este capítulo se ha realizado un análisis del principio de superposición tiempo-temperatura, empleando para ello diferentes variables relacionadas entre sí. En el Capítulo 6 se recoge el efecto de la presencia de entrecruzante en la movilidad molecular de polimetacrilatos que contienen residuos de éteres de alcoholes alifáticos. En este caso, se ha analizado el efecto de la presencia de entrecruzante tanto en los procesos de relajación secundarios, como en el proceso de relajación principal. También se llevó a cabo un análisis del efecto que la presencia de entrecruzante tiene sobre la creación de nanodominios gobernados por las cadenas laterales.
[CAT] En aquest treball es presenta un estudi de la influència de l'estructura química dels polímers en el seu comportament tèrmic, mecànic i dielèctric. Les tècniques experimentals utilitzades han sigut la calorimetria diferencial de rastreig, l'anàlisi dinamo-mecànic i l'espectroscòpia dielèctrica. Addicionalment, s'han empleat altres tècniques com la difracció de rajos X a fi de corroborar els resultats obtinguts per les primeres. En els Capítols 1 i 2 s'arreplega la introducció i els objectius, respectivament. Al Capítol 3 es presenta una breu descripció de les tècniques experimentals emprades. En el Capítol 4 es recull els resultats obtinguts en l'anàlisi comparativa de la resposta a camps de pertorbació elèctrica en un ampli rang de freqüències i temperatures de tres polimetacrilats de benzil amb dos grups metoxi en posicions 2,5-, 2,3- i 3,4-. Els resultats obtinguts assenyalen l'important efecte de la posició dels grups metoxi en l'anell aromàtic, sobre la dinàmica molecular del polimetacrilat de benzil. Els espectres obtinguts van ser molt complexos, per aquesta raó per a dur a terme un millor anàlisi es van emprar mètodes numèrics per a la transformació temps-freqüència que van incloure l'ús de tècniques de regularització paramètrica. S'ha estudiat l'efecte que el dit canvi estructural exerceix tant sobre els processos de relaxació secundària com sobre el procés de relaxació , relacionat amb la transició vítria. Així mateix, s'ha analitzat l'efecte de la posició dels grups metoxi en la formació de nanodominis en els que predominen les cadenes laterals, i el seu efecte en els processos de conducció dels materials analitzats. En el Capítol 5 s'arreplega l'estudi de la conductivitat de líquids gomosos prenent com a model el poli-(metacrilat de 2,3-dimetoxibencilo), pel seu peculiar comportament. En aquest capítol s'ha realitzat un anàlisi del principi de superposició temps-temperatura, emprant per a això diferents variables relacionades entre sí. En el Capítol 6 s'arreplega l'efecte de la presència d'entrecreuat en la mobilitat molecular de polimetacrilats que contenen residus d'èters d'alcohols alifàtics. En aquest cas, s'ha analitzat l'efecte de la presència d'entrecreuat tant en els processos de relaxació secundaris, com en el procés de relaxació principal. També es va dur a terme un anàlisi de l'efecte que la presència d'entrecreuat químic té sobre la creació de nanodominis governats per les cadenes laterals.
Carsí Rosique, M. (2015). Molecular mobility. Structure-property relationship of polymeric materials [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/59460
TESIS
Premiado
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Koukalová, Alena. « Studium lipidových membrán v nanorozlišení pomocí fluorescenční detekce jednotlivých molekul ». Doctoral thesis, 2018. http://www.nusl.cz/ntk/nusl-391388.
Texte intégralRésumé :
The complexity of cell membranes is far from being only a simple assembly of lipids and proteins separating cells from the surrounding environment. Each of the thousands of different membrane components performs its specific role in cellular functions, since a multitude of biological processes is mediated by membranes. The understanding of the molecular basis of these processes is one of the important aims of current biological research. Our research employing single- molecule fluorescence methods (e.g. FCS, FCCS, FLIM-FRET) has made a contribution to the knowledge of membrane lateral organization or mechanism of membrane fusion. Furthermore, we revealed the mechanism of membrane activity of a small natural compound. As native cell membranes are very complex structures, we performed the experiments on simplified model lipid membranes that allow studying lipid-lipid or lipid-protein interactions at the molecular level in a controlled way. The first part of this thesis deals with the mode of action of a membrane active secondary metabolite didehydroroflamycoin (DDHR). We demonstrated that DDHR is a pore-forming agent and that this activity is influenced by the presence of cholesterol. Direct visualization of intrinsic fluorescence of DDHR revealed its preferential partitioning into membrane areas...
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Schlegel, Jan. « Super-Resolution Microscopy of Sphingolipids and Protein Nanodomains ». Doctoral thesis, 2021. https://doi.org/10.25972/OPUS-22959.
Texte intégralRésumé :
The development of cellular life on earth is coupled to the formation of lipid-based biological membranes. Although many tools to analyze their biophysical properties already exist, their variety and number is still relatively small compared to the field of protein studies. One reason for this, is their small size and complex assembly into an asymmetric tightly packed lipid bilayer showing characteristics of a two-dimensional heterogenous fluid. Since membranes are capable to form dynamic, nanoscopic domains, enriched in sphingolipids and cholesterol, their detailed investigation is limited to techniques which access information below the diffraction limit of light. In this work, I aimed to extend, optimize and compare three different labeling approaches for sphingolipids and their subsequent analysis by the single-molecule localization microscopy (SMLM) technique direct stochastic optical reconstruction microscopy (dSTORM). First, I applied classical immunofluorescence by immunoglobulin G (IgG) antibody labeling to detect and quantify sphingolipid nanodomains in the plasma membrane of eukaryotic cells. I was able to identify and characterize ceramide-rich platforms (CRPs) with a size of ~ 75nm on the basal and apical membrane of different cell lines. Next, I used click-chemistry to characterize sphingolipid analogs in living and fixed cells. By using a combination of fluorescence microscopy and anisotropy experiments, I analyzed their accessibility and configuration in the plasma membrane, respectively. Azide-modified, short fatty acid side chains, were accessible to membrane impermeable dyes and localized outside the hydrophobic membrane core. In contrast, azide moieties at the end of longer fatty acid side chains were less accessible and conjugated dyes localized deeper within the plasma membrane. By introducing photo-crosslinkable diazirine groups or chemically addressable amine groups, I developed methods to improve their immobilization required for dSTORM. Finally, I harnessed the specific binding characteristics of non-toxic shiga toxin B subunits (STxBs) and cholera toxin B subunits (CTxBs) to label and quantify glycosphingolipid nanodomains in the context of Neisseria meningitidis infection. Under pyhsiological conditions, these glycosphingolipids were distributed homogenously in the plasma membrane but upon bacterial infection CTxB detectable gangliosides accumulated around invasive Neisseria meningitidis. I was able to highlight the importance of cell cycle dependent glycosphingolipid expression for the invasion process. Blocking membrane accessible sugar headgroups by pretreatment with CTxB significantly reduced the number of invasive bacteria which confirmed the importance of gangliosides for bacterial uptake into cells. Based on my results, it can be concluded that labeling of sphingolipids should be carefully optimized depending on the research question and applied microscopy technique. In particular, I was able to develop new tools and protocols which enable the characterization of sphingolipid nanodomains by dSTORM for all three labeling approachesDie Entwicklung von zellulären Lebensformen auf der Erde basiert auf der Entstehung biologischer Lipid-Membranen. Obwohl viele Techniken zur Verfügung stehen, welche es erlauben deren biophysikalische Eigenschaften zu untersuchen, sind die Möglichkeiten, verglichen mit der Analyse von Proteinen, eher eingeschränkt. Ein Grund hierfür, ist die geringe Größe von Lipiden und deren komplexe Zusammenlagerung in eine asymmetrische dicht gepackte Lipiddoppelschicht, welche sich wie eine heterogene zweidimensionale Flüssigkeit verhält. Durch die lokale Anreicherung von Sphingolipiden und Cholesterol sind Membranen in der Lage dynamische, nanoskopische Domänen auszubilden, welche lediglich mit Techniken, welche die optische Auflösungsgrenze umgehen, detailliert untersucht werden können. Ein wesentliches Ziel meiner Arbeit war es, drei Färbeverfahren für Sphingolipide zu vergleichen, erweitern und optimieren, um eine anschliessende Untersuchung mit Hilfe der einzelmolekülsensitiven Technik dSTORM (direct stochastic optical reconstruction microscopy) zu ermöglichen. Zunächst verwendete ich das klassische Färbeverfahren der Immunfluoreszenz, um Sphingolipid-Nanodomänen auf eukaryotischen Zellen mit Hilfe von Farbstoff-gekoppelten Antikörpern zu detektieren und quantifizieren. Dieses Vorgehen ermöglichte es mir, Ceramid-angereicherte Plattformen mit einer Größe von ~ 75nm auf der basalen und apikalen Membran verschiedener Zell-Linien zu identifizieren und charakterisieren. Als nächstes Verfahren verwendete ich die Klick-Chemie, um Sphingolipid-Analoge in lebenden und fixierten Zellen zu untersuchen. Eine Kombination aus Fluoreszenz-Mikroskopie und Anisotropie-Messungen erlaubte es mir Rückschlüsse über deren Zugänglichkeit und Konfiguration innerhalb der Plasmamembran zu ziehen. Hierbei lokalisierten Azid-Gruppen am Ende kurzkettiger Fettsäurereste außerhalb des hydrophoben Membrankerns, wodurch sie mittels membran-undurchlässige Farbstoffe angeklickt werden konnten. Im Gegensatz dazu, waren Azide an längeren Fettsäureresten weniger zugänglich und konjugierte Farbstoffe tauchten tiefer in die Plasmamembran ein. Durch die Einführung photoreaktiver Diazirin-Gruppen oder chemisch modifzierbarer Amin-Gruppen wurden Wege geschaffen, welche eine Immobilisierung und anschließende Analyse mit Hilfe von dSTORM ermöglichen. Schließlich nutzte ich das spezifische Bindeverhalten der nicht toxischen B Untereinheiten von Shiga- (STxB) und Cholera-Toxin (CTxB) aus, um Glycosphingolipid Nanodomänen im Kontext einer Neisseria meningitidis Infektion zu untersuchen. Unter physiologischen Bedingungen waren diese homogen in der Plasmamembran verteilt, jedoch reicherten sich CTxB-detektierbare Ganglioside um eindringende Bakterien an. Darüber hinaus konnte ich einen Zusammenhang zwischen der zellzyklusabhängigen Expression von Glycosphingolipiden und dem Eindringen der Bakterien herstellen. Eine Absättigung der Zucker an der äußeren Membran durch CTxB-Vorbehandlung reduzierte die Anzahl von invasiven Bakterien signifikant und bestätigte die Schlüsselrolle von Gangliosiden bei der Aufnahme von Bakterien. Meine Ergebnisse legen Nahe, dass das Färbeverfahren für Sphingolipide an die jeweilige Fragestellung und Mikroskopietechnik angepasst werden sollte. Im Rahmen dieser Arbeit konnten neue Werkzeuge und Protokolle geschaffen werden, die die Charakterisierung von Sphingolipid-Nanodomänen mittels dSTORM für alle drei Färbeverfahren ermöglichen
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Jones, Joseph F. « Examining initial bacterial adhesion oriented adhesion and surface nanodomains / ». 2005. http://www.etda.libraries.psu.edu/theses/approved/WorldWideIndex/ETD-1107/index.html.
Texte intégral
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« Photoalignment Control of Nanodomains in Liquid Crystalline Block Copolymer Thin Films ». Thesis, 2007. http://hdl.handle.net/2237/8484.
Texte intégral
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MORIKAWA, Yuichi, et 雄市 森川. « Photoalignment Control of Nanodomains in Liquid Crystalline Block Copolymer Thin Films ». Thesis, 2007. http://hdl.handle.net/2237/8484.
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Šachl, Radek. « Lokalizace fluorescenčních značek a určování velikostí lipidových nanodomén pomocí moderních fluorescenčních metod ». Doctoral thesis, 2012. http://www.nusl.cz/ntk/nusl-308511.
Texte intégralRésumé :
iii Organizations Umeå University Department of Chemistry, Umeå, SE & Charles University in Prague, Faculty of Science, Prague, CZ Document name Doctoral thesis Date of issue February 2012 Author Radek Šachl Title Localisation of Fluorescent Probes and the Estimation of Lipid Nanodomain Sizes by Modern Fluorescence Techniques Abstract The thesis is divided into two major parts. The first part focuses on the localisation of probes in lipid/polymeric bilayers and in GM1 micelles. Included in this thesis is a new approach based on electronic energy transfer/migration (FRET/DDEM), which efficiently determines transversal positions of fluorescent molecules in lipid bilayers. This approach has been used to locate newly synthesized lipid probes in DOPC bilayers. The label was introduced at the end of sn-2 acyl chains of variable length. Analytical models accounting for FRET exist for a limited number of basic geometries. Here, a combination of FRET and Monte Carlo simulations enables the localisation of probes in bicelles and in bilayers containing pores, i.e. in lipid systems with variable curvature, or in non-homogenous lipid systems. This approach has been used to test whether conical-like fluorescence probes have an increased affinity to highly curved regions, which would enable preferential labelling of...
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Gao, Yan. « Nanodomain Structure and Energetics of Carbon Rich SiCN and SiBCN Polymer-Derived Ceramics ». Phd thesis, 2014. https://tuprints.ulb.tu-darmstadt.de/3736/1/Yan%20Gao%2C%20Ph.D.Thesis%202014.pdf.
Texte intégralRésumé :
This Ph.D. thesis focuses on the synthesis, processing, solid state structure, nanodomain structure, structural evolution, thermodynamic stability, and functional properties of carbon rich SiCN and SiBCN ceramics derived from preceramic polymers with tailored compositions and structures. The main objective of the studies is to better understand the effects of the composition and structure of the starting precursors, on the behavior of the resultant ceramics.
First, a set of preceramic polymers with systematically varied compositions and structures were synthesized. They are linear polysilylcarbodiimides and polysilazanes, and their boron modified counterparts; branched polysilsesquicarbodiimide and its 13C/15N isotope-enriched counterpart; and branched polysilsesquiazane. The synthesis of these polymers was investigated using NMR, FT-IR, Raman and TG/DTG. The results demonstrate that the obtained precursors exhibit the expected compositions and structures.
Then, the effects of processing route on the thermal stability of PDCs were studied by comparing bulk ceramics with their powder counterparts. The thermal transformation was investigated using FT-IR, Raman spectroscopy, XRD, TG/DTG and TEM. The results reveal that bulk ceramics are more thermally stable than their powder counterparts in terms of resistance to crystallization and decomposition. The Si(B)CN ceramic powders derived from poly(boro)phenylvinylsilylcarbodiimide contain α/β-SiC crystallites when heat treated at 1400ºC, while their bulk counterparts prepared at the same temperature remain completely amorphous. It is also found that bulk ceramics exhibit less weight loss than their powder analogues at temperatures up to 2100°C, especially in the case of SiCN bulk ceramics. The higher thermal stability of bulk ceramics as compared with the powder counterparts is likely due to the powders have greater surface area which enhances the carbothermal reaction and silicon carbide crystallization. In addition, the boron modification impedes the degradation of silicon nitride in both bulk and powder samples, similar to previously reported results.
Next, the energetics of PDCs was investigated using high temperature oxidative drop solution calorimetry on (i) ceramics derived from branched and linear polysilylcarbodiimide; (ii) ceramics derived from poly(boro)silazanes pyrolyzed at different temperatures. The results reveal that the ceramics derived from the branched polymer is energetically more stable than those from the linear polymer. Structural analysis using MAS NMR suggests that the increased energetic stability of the ceramics derived from the branched polymer is likely due to the presence of hydrogen at the mixed bonding environments consisting of N, C and Si atoms. These environments make up the interfacial region between the Si3N4 and “free” carbon nanodomains. For both the linear and branched polymers, the ceramics derived at 800 ºC are energetically more stable than those derived at 1100 ºC. The study of group (ii) reveals the effect of the pyrolysis temperature on the structural evolution and energetics of poly(boro)silazane-derived Si(B)CN ceramics. These ceramics contain mixed SiCxN4-x(x=0-4) tetrahedra. MAS NMR spectroscopy of the 1100ºC and 1400ºC ceramics reveals that the structural evolution involves the following processes: (i) the demixing of SiCxN4-x mixed bonding environments, (ii) the cleavage of mixed bonds at the interdomain regions, and (iii) the coarsening of domains. Calorimetry results demonstrate that this structural evolution is favorable in both enthalpy and free energy.
We also investigated the solid state structures and nanodomain structures of the ceramics derived from the tailored polymers. The MAS NMR results indicate that the SiCN ceramics derived from polyphenylvinylsilylcarbodiimide and polymethylvinylsilylcarbo- diimide both contain nanodomains of silicon nitride and “free” carbon. In the ceramics prepared from the phenyl-containing polymer, the “free” carbon and silicon nitride domains are basically isolated, while in the ceramics derived from the methyl-containing polymer, “free” carbon and silicon nitride domains are connected via C-N bonds. The SiBCN ceramics derived from boron-modified polysilylcarbodiimides have an additional B-containing phase which is located at the interface between the silicon nitride and the “free” carbon phase. On the other hand, the SiCN ceramics derived from polysilazanes contain “free” carbon and mixed bonded SiCxN4-x(x=0-4) nanodomains. The SiCxN4-x (x=0-4) domain consists of a core of SiN4 tetrahedra, which connect to “free” carbon domain via SiN3C, SiN2C2, SiNC3, and SiC4 tetrahedra. SiC4 tetrahedra make up the most outer shell of mixed bonded SiCxN4-x(x=0-4) nanodomains. The SiBCN ceramics derived from boron-modified polysilazanes have an additional B-containing phase which connects the SiC4 with the “free” carbon domains. SAXS results indicate that (i) the size of Si-containing nanodomains in SiCN ceramic is larger than in their SiBCN counterparts; (ii) the ceramics derived from phenyl-containing polymers exhibit smaller Si-containing nanodomains than those derived from methyl-containing polymers; and (iii) the size of Si-containing domains increases with pyrolysis temperature in all ceramics.
Finally, some functional behaviors of the resultant ceramics were investigated. The electrochemical properties of the ceramics were investigated to explore their applicability as anode materials for lithium-ion batteries. The results reveal that the SiCN ceramics derived from both linear and branched polymers are suitable anode materials for lithium-ion batteries. In particular, the SiCN ceramics derived from linear polyphenylvinylsilazane demonstrate outstanding performance in terms of cycling stability and capacity at high currents. In addition, the AC conductivity was characterized using impedance spectroscopy. It is found that the impedance spectra vary in accordance to the different microstructures of the ceramics, which, in turn, are related to the chemistry of the precursors. Conduction in SiCN ceramics is dominated via “free” carbon and SiC phases in series, as analyzed by two semicircles in the Nyquist plot. Conduction in SiBCN ceramics is dominated by one phase, and is thus represented by one semicircle in the Nyquist plot. Nonconducting BN forms the interfacial phase between the “free” carbon and SiC phases, and isolates the discontinuous phase from the continuous phase. Therefore, conduction in SiBCN ceramics is dominated by the continuous phase, whether it is “free carbon” or SiC.
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Ross, Elizabeth Ina. « Tuning the electronic and molecular structures of catalytic active sites with oxide nanodomains ». 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3314501.
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Zandt, Maximilian. « Nanodomain clustering mechanisms of Junctophilin-2 in human kidney, cardiac and skeletal muscle cells ». Doctoral thesis, 2020. http://hdl.handle.net/21.11130/00-1735-0000-0005-1475-1.
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Marquês, Joaquim Trigo. « Supported lipid bilayers with micro/nanodomains in the study of membrane lipid organization and interactions ». Doctoral thesis, 2015. http://hdl.handle.net/10451/22728.
Texte intégralRésumé :
Tese de doutoramento, Química (Bioquímica), Universidade de Lisboa, Faculdade de Ciências, 2015The lateral organization of lipid bilayers into domains, such as lipid rafts or gel domains, and how they influence the properties and function of membranes is a current topic in biophysics. In this work, the properties of single and multicomponent lipid bilayers (whether supported on a solid substrate or free in solution as liposomes) exhibiting different phase behavior were studied by a wide range of characterization techniques – atomic force microscopy, ellipsometry, cyclic voltammetry, quartz crystal microbalance, surface plasmon resonance and fluorescence spectroscopy. The phase diagram for the binary mixture 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) / N-octadecanoyl(2S,3S,4R)-2-amino-1,3,4-octadecanetriol (Phytoceramide (PhyCer)) is proposed by combining data from different techniques. The formation of complexes between POPC and PhyCer with properties similar to those found for gel domains in vivo is anticipated. These POPC/PhyCer complexes occur at two distinct stoichiometries, 3:1 and 1:2. The influence of membrane lateral organization was investigated in the context of the interaction of small molecules with lipid bilayers, namely ethanol and epinephrine. The effects of ethanol were studied in supported lipid bilayers (SLB) formed on mica spanning a phase behavior from single fluid to liquid ordered (lo) / liquid disordered (ld) phase coexistence. It was concluded that the lateral organization of lipids into domains, but not the specific lipid composition, plays a determinant role on the effects induced by ethanol. Another purpose of this work was to study the properties of supported lipid bilayers (SLB) formed on gold surfaces. For the first time raft-containing SLB were formed directly on bare gold and the substrate seems to influence the properties of the lipid bilayer since an unexpected proportion of lipid domains was obtained and corrugations were observed in the liquid disordered phase. A lipid-based biosensing interface consisting on a SLB formed on top of a L-cysteine-modified gold was also developed for the detection of membrane-interacting electroactive molecules.The interaction of epinephrine, an electroactive molecule, was evaluated using both liposomes and SLB formed on Lcysteine-modified gold. Despite the weak interaction between epinephrine and liposomes as determined by fluorescence spectroscopy, its redox signal could be clearly detected by cyclic voltammetry when adsorbed on SLB of various compositions. Moreover, voltammetric data allowed to estimate a membrane/water partition coefficient for epinephrine. The results presented show how lateral membrane organization and composition may influence its function and properties.
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Mohamedgamil, Sana Siddig Abdelrahman. « Organization and dynamics of class C GPCR nanodomains in neurons visualized by single-molecule microscopy ». Doctoral thesis, 2020. https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-178963.
Texte intégralRésumé :
Despite the large number of G protein-coupled receptors (GPCRs) expressed in the central nervous system (CNS), little is known about their location, organization, and dynamics in functional nanodomains at synapses. Class C GPCRs including metabotropic glutamate receptors (mGluRs) and the γ-aminobutyric acid subtype B receptor (GABABR) mediate several key functions in synaptic transmission. However, it is still insufficiently understood how these receptors function at synapses to modulate neurotransmission. One limitation is the availability of techniques to examine receptors with high spatiotemporal resolution in physiologically relevant cells. To investigate the distribution and spatiotemporal dynamics of mGluR4 and GABABR in cerebellar slices and cultured hippocampal neurons, I used advanced imaging techniques, including single-molecule imaging and superresolution microscopy with high spatial (10-20 nm) and temporal (20 ms) resolution. The presynaptic active zone (AZ) is a highly organized structure that specializes in neurotransmitter release. mGluR4 is a prototypical presynaptic class C GPCR. mGluR4 mediates an inhibitory effect on presynaptic glutamate release mainly via the inhibition of P/Q type voltage dependent calcium channels (CaV2.1). In this study, I analyzed the organization of mGluR4 at the synapse between parallel fibers and Purkinje cells in the mouse cerebellum with near-molecular resolution using two-color direct stochastic optical reconstruction microscopy (dSTORM). Quantitative analyses revealed a four-fold mGluR4 enrichment at parallel fiber AZs. I found that an AZ contains 29 mGluR4 nanoclusters on average. Each nanocluster contains one or two mGluR4s, with few nanoclusters containing three or more receptors. To assess the spatial distribution of mGluR4 relative to functional active zone elements such as CaV2.1 and Munc 18-1 (an essential component of the synaptic secretory machinery), a distance-based colocalization analysis was used. The analysis revealed positive correlation between mGluR4 and both proteins at a distance of 40 nm. Interestingly, mGluR4 showed a higher positive correlation to Munc 18-1 in comparison to CaV2.1. These results suggest that mGluR4 might directly inhibit the exocytotic machinery to reduce glutamate release from the synaptic vesicles in addition to its role in the inhibition of presynaptic calcium influx. The revealed high degree of mGluR4 organization may provide a new ultrastructural basis to explain the depressive effect of mGluR4 on the neurotransmission. Moreover, I directly imaged GABABR dynamic behavior with high spatiotemporal resolution in living hippocampal neurons utilizing single-molecule total internal reflection fluorescence microscopy (TIRFM). To this purpose, the GABAB1 subunit was engineered with an N-terminal SNAP-tag to enable specific labeling with bright organic fluorophores. On the plasma membrane surface, immobile and mobile GABABRs were detected at both synaptic and extrasynaptic compartments. A mean square displacement analysis (MSD) revealed characteristic dynamic patterns of GABABR depending on receptor location inside or outside of the synapses. The majority of receptors belonging to the extrasynaptic pool displayed rapid and free diffusion. In contrast, approximately 80% of receptors residing at the synaptic compartments were immobile or confined within limited regions. Receptors located at pre- and post-synaptic sites showed a similar behavior. GABABR lateral diffusion patterns inside and outside synapses might be important for the regulation of efficacy of synaptic inhibition. Altogether, this study puts forward previously unknown GPCR nanoscopic details in functional nanodomains. GPCR spatial organization might be important for the efficiency, fidelity, and rapid signaling required for synaptic transmissionTrotz der großen Anzahl an G Protein-gekoppelten Rezeptoren (GPCRs) die im zentralen Nervensystem (ZNS) exprimiert werden, ist deren Lokalisierung, Anordnung und Dynamik in funktionellen Nanodomänen an Synapsen gegenwärtig weitgehend unbekannt. Klasse C GPCRs, einschließlich metabotroper Glutamatrezeptoren (mGluRs) und des γ- Aminobuttersäure-B-Rezeptors (GABABR), vermitteln einige Schlüsselfunktionen der synaptischen Übertragung. Grundlegende Prinzipien wie diese Rezeptoren an Synapsen funktionieren, um die Neurotransmission zu modulieren, sind jedoch nur unvollständig verstanden. Eine Schwierigkeit ist die Verfügbarkeit von Techniken zur Untersuchung von Rezeptoren mit hoher raum-zeitlicher Auflösung in physiologisch relevanten Zellen. Um die Anordnung und die raum-zeitliche Dynamik von mGluR4 und GABABR in Kleinhirnschnitten und kultivierten hippocampalen Neuronen zu untersuchen, verwendete ich neue optische Verfahren wie Einzelmolekül- und hochauflösende Mikroskopie (dSTORM) mit hoher räumlicher (10-20 nm) und zeitlicher Auflösung (20 ms). Die präsynaptische aktive Zone (AZ) ist eine hoch organisierte Struktur, die auf die Transmitterausschüttung spezialisiert ist. Der mGluR4 ist ein prototypischer, präsynaptischer Klasse C GPCR. Hauptsächlich durch die Inhibierung spannungsgesteuerter Calciumkanäle des P/Q-Typs (CaV2.1) vermittelt mGluR4 eine inhibitorische Wirkung auf die Glutamatfreisetzung.In dieser Arbeit analysierte ich die Organisation des mGluR4 an der Synapse zwischen parallelen Fasern und Purkinje-Zellen im Kleinhirn der Maus unter Verwendung der Zweifarben direkten stochastischen optischen Rekonstruktionsmikroskopie (dSTORM). Quantitative Analysen zeigten eine vierfache höhere Anreicherung von mGluR4 an den Parallelfaser-AZs. Ich fand heraus, dass eine AZ im Durchschnitt 29 mGluR4- Nanocluster enthält. Jeder Nanocluster enthält ein oder zwei mGluR4s, wobei wenige Nanocluster drei oder mehr Rezeptoren enthalten. Um die räumliche Verteilung von mGluR4 relativ zu funktionellen Elementen aktiver Zonen, wie CaV2.1 und Munc 18-1( ein wichtiges Protein der exozytotischen Maschinerie), zu bestimmen, wurde die Abstandsbasierte-Co- Lokalisierung-Analyse verwendet. Co-Lokalisierung wurde zwischen mGluR4 und beiden Proteinen in einem Abstand von 40 nm detektiert. Interessanterweise wurde für Munc 18-1 eine höhere positive Korrelation zu mGluR4 identifiziert. Dies deutet darauf hin, dass mGluR4, zusätzlich zu seiner Rolle bei der Hemmung des präsynaptischen Calciumeinstroms, die exozytotische Maschinerie zur Verringerung der Freisetzung von Glutamat aus sekretorischen Organellen direkt hemmen könnte. Der gezeigte hohe Grad der mGluR4-Organisation könnte eine neue ultrastrukturelle Grundlage zur Erklärung der depressiven Wirkung von mGluR4 auf die synaptische Übertragung liefern. Außerdem habe ich das dynamische Verhalten von GABABR direkt mit hoher räumlicher und zeitlicher Auflösung in lebenden hippocampalen Neuronen durch Einzelmolekül-TIRFM visualisiert. Zu diesem Zweck wurde die GABAB1-Untereinheit mit einem N-terminalen SNAP- Tag konstruiert, um eine spezifische Markierung mit hellen organischen Fluorophoren zu ermöglichen. Auf der Oberfläche der Plasmamembran wurden immobile und mobile GABABRs in synaptischen und extrasynaptischen Kompartimenten nachgewiesen. Die mittlere quadratische Verschiebung (mean square displacment analysis (MSD)) zeigte charakteristische dynamische Muster von GABABR in Abhängigkeit der Position der Rezeptoren innerhalb oder außerhalb der Synapsen. Die Mehrheit der Rezeptoren im extrasynaptischen Pool zeigte schnelle und freie Diffusion. Im Gegensatz dazu waren ungefähr 80% der synaptischen Rezeptoren immobile oder auf begrenzte Regionen beschränkt. Rezeptoren an prä- und postsynaptischen Stellen zeigten ein ähnliches Verhalten. GABABR- Diffusionsmuster innerhalb und außerhalb von Synapsen könnten außerdem für die Regulierung der Wirksamkeit der synaptischen Hemmung von Bedeutung sein. Insgesamt zeigen diese Ergebnisse bisher unbekannte Erkenntnisse zu nanoskopischen Einzelheiten von GPCRs in funktionellen Nanodomänen. Die räumliche Organisation von GPCR kann für die Effizienz, Genauigkeit und schnelle Signalisierung der Neurotransmission wichtig sein
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Huang, Wen-Yu, et 黃文昱. « Revealing Nanodomains of Bulk-heterojunction P3HT:PCBM Organic Solar Cell with Scattering-type Scanning Near-field Optical Microscopy ». Thesis, 2018. http://ndltd.ncl.edu.tw/handle/u5xjjb.
Texte intégralRésumé :
碩士國立臺灣大學
物理學研究所
106
Polymer-based organic solar cell has been seen as a potential candidate of next-generation photovoltaics. The introduction of bulk-heterojunction (BHJ) concept — a blended film of electron donor and acceptor that forms an interconnected network — is the key to greatly improve the power conversion efficiency (PCE). The domain size in the blended layer of donor and acceptor should be comparable to the exciton diffusion length (~20 nm) for best device performance. Revealing the relationship between nanostructure morphology to the electro-optical properties of organic solar cell device therefore requires a characterization tool with spatial resolution down to less 10 nanometers. In this study, nanostructure morphology of the pristine P3HT and the blended P3HT:PCBM films—a prototypical polymer-based organic solar cell was investigated by a heterodyne-based scattering-type scanning near-field optical microscopy (s-SNOM) with an excitation wavelength of 632.8 nm. The obtained s-SNOM images were interpreted with point-dipole and modified point dipole models. Theoretical prediction based on these two models show that the near-field amplitude and phase signals of P3HT are larger than that of PCBM, while these two near-field signals of ordered P3HT are larger than that of disordered P3HT. Based on these two near-field traits, the s-SNOM image of pristine P3HT displays nanometer-scaled connected network of ordered P3HT, while that of blended P3HT:PCBM shows isolated nanodomains of P3HT. Last but not least, the portrayed area ratio between P3HT and PCBM is smaller than the mixing ratio, indicating that the actual distribution of P3HT domains is not uniform over the whole active layer. This study demonstrates the capability of s-SNOM as a tool in identifying nanostructure morphology of blended P3HT:PCBM film in BHJ polymer solar cells.
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Lee, Seok-Jin. « Spatiotemporal Dynamics of Calcium/calmodulin-dependent Kinase II in Single Dendritic Spines During Synaptic Plasticity ». Diss., 2011. http://hdl.handle.net/10161/3818.
Texte intégralRésumé :
Synaptic plasticity is the leading candidate for the cellular/molecular basis of learning and memory. One of the key molecules involved in synaptic plasticity is Calcium/calmodulin-dependent Kinase II (CaMKII). Synaptic plasticity can be expressed at a single dendritic spine independent of its neighboring dendritic spines. Here, we investigated how long the activity of CaMKII lasts during synaptic plasticity of single dendritic spines. We found that CaMKII activity lasted ~2 minutes during synaptic plasticity and was restricted to the dendritic spines undergoing synaptic plasticity while nearby dendritic spines did not show any change in the level of CaMKII activity. Our experimental data argue against the persistent activation of CaMKII in dendritic spines undergoing synaptic plasticity and suggest that the activity of CaMKII is a spine-specific biochemical signal necessary for synapse-specificity of synaptic plasticity. We provide a biophysical explanation of how spine-specific CaMKII activation can be achieved during synaptic plasticity. We also found that CaMKII is activated by highly localized calcium influx in the proximity of Voltage-dependent Calcium Channels (VDCCs) and a different set of VDCCs and their respective Ca2+ nanodomains are responsible for the differential activation of CaMKII between dendritic spines and dendritic shafts.
Dissertation
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Kirsch, Wolfgang [Verfasser]. « Biophysical analysis of the spatial and temporal distribution of free Ca2+ ions within micro- and nanodomains of muscle cells / presented by Wolfgang Kirsch ». 2000. http://d-nb.info/961763817/34.
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« Live-cell transforms between calcium transients and FRET responses for troponin C-based calcium sensors : Their application to probe calcium in the calcium channel nanodomain ». THE JOHNS HOPKINS UNIVERSITY, 2008. http://pqdtopen.proquest.com/#viewpdf?dispub=3309816.
Texte intégral
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Li, Chung-Ping, et 李中斌. « Collective Electron Transport in Au and CdSe Nanoparticles Self-Assembled in the Poly(4-vinylpyridine) Nanodomains of a Poly(styrene-b-4-vinylpyridine) Diblock Copolymer Thin Film ». Thesis, 2006. http://ndltd.ncl.edu.tw/handle/82987311338008251080.
Texte intégralRésumé :
博士國立交通大學
材料科學與工程系所
94
In this dissertation, thin films that consisted of Au and CdSe nanoparticles (NPs), self-assembled in a poly(4-vinylpyridine) (P4VP) nanodomain of poly(styrene-b-4-vinylpyridine) (PS-b-P4VP) diblock copolymer were prepared by polar interaction and solvent selectivity. Collective electron transport of these organic nanodomain confined nanoparticles were investigated. From conductive atomic force microscopy and device measurements, we found that the electron tunneling rate constant in the case of CdSe quantum dots confined in a P4VP nanodomain is much larger than that in the randomly-distributed case. The calculated electron tunneling coefficient for hopping between confined CdSe quantum dots was 0.3 1/Å. The conductivity of the CdSe/P4VP nanodomain increased upon increasing the amount of CdSe, following a percolation model (Chapter 2). From the current–voltage characteristics of PS-b-P4VP thin films that consist of Au NPs, we found that the collective electron transport behavior of Au NPs sequestered in the spherical P4VP nanodomains was dictated by Coulomb blockade and was quasi one-dimensional, as opposed to the three-dimensional behavior displayed by Au NPs that had been dispersed randomly in homo-P4VP (Chapter 3). Moreover, CdSe nanorods self-assembled in the P4VP nanodomains of a PS-b-P4VP diblock copolymer thin film were aligned under the influence of the polarization forces created by an applied electric field. The electron mobilities of CdSe/P4VP nanodomains incorporating the out-of-plane CdSe nanorods were much larger than those incorporating in-plane nanorods (chapter 4).
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Saka, Kırlı Sinem. « Studying Protein Organization in Cellular Membranes by High-Resolution Microscopy ». Doctoral thesis, 2013. http://hdl.handle.net/11858/00-1735-0000-0023-98FB-0.
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