Rozprawy doktorskie: „Lipidic model membranes”

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Oldham, Alexis Jean. "Modulation of lipid domain formation in mixed model systems by proteins and peptides". View electronic thesis, 2008. http://dl.uncw.edu/etd/2008-1/r1/oldhama/alexisoldham.pdf.

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Azouz, Mehdi. "Alzheimer's disease neurotoxic peptides : towards a comprehension of their modes of action on model membranes". Thesis, Bordeaux, 2019. http://www.theses.fr/2019BORD0419.

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La maladie d'Alzheimer (MA) est une neuropathologie complexe qui constitue la principale forme de démence chez l'être humain. Étroitement associée au vieillissement, elle se manifeste par une perte progressive de la mémoire et des fonctions cognitives. Avec 30 millions d’individus concernés au niveau mondial et des estimations voyant ce chiffre quadrupler d'ici 2050, elle constitue aujourd’hui une menace sociétale majeure. L’atrophie cérébrale observée chez les patients atteints de la MA est la conséquence d’un long processus de neurodégénérescence qui intervient au niveau moléculaire et s’amorce bien avant l’apparition des symptômes. Deux marqueurs histopathologiques ont été identifiés comme étant associés à ce processus : les plaques séniles, composées du peptide Abêta et les dégénérescences neurofibrillaires constituées de la protéine Tau. Ces deux molécules, considérées comme les protagonistes décisifs du développement de la MA, concentrent les recherches afin de mieux comprendre leurs rôles dans le processus neurodégénératif et pouvoir mettre en place des solutions thérapeutiques, inexistantes à ce jour.Un des axes de recherche majeurs se focalise sur l’interaction de ces peptides avec la membrane plasmique. L’occurrence d’un tel phénomène pourrait potentiellement être en cause dans la mort neuronale s’il s’avérait délétère. Il est donc capital d’étudier en détail ces processus afin d’identifier les facteurs qui pourraient conduire Abêta et Tau à endommager l’intégrité des membranes. De nombreux travaux ont démontré que certains lipides pouvaient promouvoir ces interactions. Cependant, les conclusions sont parfois divergentes et un consensus commun reste à trouver quant à leurs rôles.Ce travail de thèse s’est consacré à l’étude des modes d’action du peptide Abêta et d’un fragment clé de la protéine Tau, le peptide K18, sur des membranes modèles, en se focalisant principalement sur l’influence de certains lipides. Afin d’élucider les mécanismes qui régissent ces phénomènes, les processus de solubilisation membranaires ont dans un premier temps été étudiés avec des molécules amphiphiles bien caractérisées : les détergents. Cette étude a permis d’établir que les phénomènes de solubilisation membranaires peuvent varier en fonction de la composition membranaire et démontrer de la sélectivité.Le cœur du projet était de visualiser les effets des peptides amyloïdes Abêta et K18 sur des modèles membranaires, les bicouches supportées, avec pour principale technique d’investigation la microscopie à force atomique. Elle nous a permis d’observer ces phénomènes in situ, en conditions physiologiques et à l’échelle sub-micrométrique. Nous avons pu montrer que la composition membranaire était un facteur pouvant moduler l’interaction avec Abêta. L'étude établit que les domaines lipidiques favorisent les perturbations membranaires induites par le peptide. Il est proposé que des défauts d'empilement lipidiques aux interfaces de ces domaines agissent comme des sites d'adsorption du peptide, menant à la destruction des membranes. En utilisant la même approche, avec des compositions lipidiques plus en adéquation avec Tau, nous avons pu établir que K18 induisait également des effets de solubilisation en fonction de la nature des lipides dans la membrane et des propriétés qui leurs sont associées.Dans les deux cas, nous montrons que les effets délétères que peuvent induire ces peptides se manifestent par des effets comparables à ceux des détergents et sont dépendants de la composition des membranes. L’agrégation des peptides, qui peut conduire à leur fibrillation, n’a également été mise en évidence qu’en présence de certains lipides.Ce travail de thèse apporte de nouvelles informations sur le caractère décisif de la membrane à pouvoir moduler les interactions avec les peptides Abêta; et K18. Par extension aux membranes cellulaires, ces phénomènes pourraient potentiellement être associés aux processus neurodégénératifs impliqués dans la MA
Alzheimer’s disease is a complex neuropathological disorder that constitutes the prime form of dementia. Intimately related to ageing, it is associated to the gradual loss of memory and cognitive functions in individual suffering from the pathology. With nearly 30 million people concerned today, and the alarming trends predicting this figure to increase fourfold by 2050, Alzheimer’s disease will constitute a major burden for our societies in the upcoming decades. The cerebral atrophy occurring within the brain results from slow and progressive neurodegenerative mechanisms triggered many years before the appearance of the first symptoms. Two histopathological markers have been identified as strongly associated to the neurodegeneration: the senile plaques, majorly composed of the amyloid peptide Abeta, and the neurofibrillary tangles, constituted of the abnormally phosphorylated form of Tau protein. These two molecules, hence considered as the main culprits of the disease, are therefore under the spotlight of researchers who try to better understand the respective roles in the neurodegeneration process and uncover therapeutic solutions to a still uncurable disease.One of the promising research axis is focusing on the interplay between these molecules and the plasma membrane as potential interactions could convincingly rationalize the neural cell deaths if they happened to be deleterious. Therefore, investigate these interactions in detail is of primary importance to identify the factors that might drive Abeta and Tau to cause damages on membranes. A strong body of evidences has demonstrated that certain lipids could promote these interactions and are then suspected to be involved into detrimental phenomena. However, numerous results appear to be contradicting and consensual conclusions are still lacking.This PhD was dedicated to the investigation of the effects of Abeta and K18, a key peptide fragment of Tau protein, on membranes with a particular focus on the influence of lipids. The aim of this work was to elucidate the action mechanisms of these peptides.To first comprehend how membrane damages can be induced, we first focused on the solubilising ability of extensively used amphiphile agents: detergents. As a first study, we revealed that the membrane composition and the physicochemical properties of lipids play an important role in driving the solubilisation of the bilayer, a process that can even lead to a selectivity during the lipid extraction.The core part of the project was to visualize the effects of the amyloid peptides Abeta and K18 on supported lipid bilayers as membrane models, using atomic force microscopy as an investigation technique. With its high spatial resolution and its ability to operate in physiological milieu, this approach has shown that the membrane composition could promote membrane disruption induced by Abeta oligomers in a lipid-dependent manner. More importantly, we propose that packing defects at the interface of membrane domains act as adsorption and nucleation sites leading to membrane damages.Using the same strategy, we observed that K18 could also induce solubilisation phenomenon and demonstrated to be sensitive to the aspect of lipid order in membranes.In both cases, we highlighted that these peptides could be detrimental to supported lipid bilayers and that their disruptive abilities, associated to detergent-like mechanisms, were intimately dependent of lipids. We also show that the aggregation, a phenomenon that can lead to the peptides fibrillation can only be triggered in presence of certain lipids.This work provides important insights about the decisive role of membrane composition in modulating interactions with the Abeta and K18. This interplay could constitute one of the numerous factors that promote neurotoxic phenomena, taking part in the complex neurodegenerative processes associated to Alzheimer’s disease

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Ury-Thiery, Vicky. "Agrégation in vitro de la protéine amyloïde Tau et étude de son impact sur des modèles membranaires par différentes méthodes biophysiques". Electronic Thesis or Diss., Bordeaux, 2024. http://www.theses.fr/2024BORD0440.

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Les maladies neurodégénératives, telles que les maladies d'Alzheimer et de Parkinson, affectent les fonctions cognitives et motrices. Elles se caractérisent par une perte progressive de neurones, sans possibilité de régénération. Avec le vieillissement de la population, ces pathologies, principalement liées à l'âge, représentent un enjeu sociétal majeur. L'absence de diagnostic précoce, de traitements efficaces et la méconnaissance des mécanismes en jeu soulignent la nécessité d'en approfondir la compréhension. Les patients atteints de ces maladies présentent des accumulations de protéines anormales sous forme d’agrégats insolubles, dans ou à proximité des cellules cérébrales. Bien que chaque protéinopathie présente des agrégats spécifiques, elles partagent des caractéristiques communes, notamment leur structure appelée amyloïde. Ces amyloïdes, formés par l’auto-assemblage de monomères protéiques mal conformés par empilement, adoptent une structure caractéristique dite en cross-β. Plusieurs protéines amyloïdes pathogènes ont été identifiées et sont associées à diverses maladies neurodégénératives. La protéine Tau, impliquée dans la maladie d'Alzheimer et plus largement dans un groupe de démences appelées tauopathies, est principalement localisée dans les neurones, où elle stabilise les microtubules, éléments structurants du cytosquelette cellulaire. Toutefois, dans des conditions pathologiques, Tau se dissocie des microtubules, devient hyperphosphorylée et forme des agrégats amyloïdes fibrillaires. Les mécanismes exacts de cette agrégation restent mal compris. L'étude de l'agrégation de Tau repose sur la production in vitro de fibres amyloïdes. En raison de sa solubilité élevée liée à sa charge positive, la formation de ces fibres nécessite l’ajout de molécules polyanioniques, appelées cofacteurs, telles que l’héparine (un polysaccharide), des ARN ou des lipides. Cependant, des incertitudes demeurent quant au rôle précis de ces cofacteurs : catalysent-ils simplement l'agrégation ou sont-ils intégrés dans la structure des fibres ? Si tel est le cas, quel impact cela a-t-il sur la morphologie des agrégats ? La capacité de Tau à s'agréger en présence de lipides suscite des interrogations sur son comportement vis à vis des différentes membranes des neurones. L’interaction de Tau avec les membranes plasmiques a été démontrée, et pourrait jouer un rôle autant dans des processus physiologiques que pathologiques. Tau, en présence de lipides anioniques, altère-t-elle l’intégrité membranaire ? Qu'en est-il des lipides non anioniques ? Pour répondre à ces questions, ce projet de thèse combine plusieurs approches biophysiques : spectroscopie infrarouge à réflexion totale atténuée (ATR-FTIR), microscopie à force atomique (AFM), microscopie électronique à transmission (MET) et résonance plasmonique de surface par ondes guidées (PWR). L’étude est structurée autour de deux axes principaux : (i) caractériser l’agrégation de Tau en présence de différents cofacteurs anioniques (héparine, ARN, phospholipides) et étudier l'impact sur la morphologie des fibres ; (ii) évaluer l’effet de l’interaction de Tau avec des membranes lipidiques de différentes compositions sur leur intégrité. Les résultats de cette thèse apportent de nouvelles perspectives sur les mécanismes pathogéniques de Tau et pourraient contribuer à une meilleure compréhension des tauopathies, ainsi qu'au développement de stratégies thérapeutiques
Neurodegenerative diseases, such as Alzheimer’s and Parkinson’s, affect cognitive and motor functions. They are characterized by a progressive loss of neurons, with no possibility of regeneration. With an aging population, these predominantly age-related diseases represent a major societal challenge. The lack of early diagnosis, effective treatments, and understanding of the underlying mechanisms highlights the need for further investigation. Patients suffering from these diseases exhibit abnormal protein accumulations in the form of insoluble aggregates, within or near brain cells. Although each proteinopathy presents specific aggregates, they share common features, notably their amyloid structure. These amyloids, formed by the misfolded protein monomers’ self-assembly through stacking, adopt a characteristic cross-β structure. Several pathogenic amyloid proteins have been identified and are associated with various neurodegenerative diseases. The Tau protein, implicated in Alzheimer’s disease and more broadly in a group of dementias known as tauopathies, is primarily located in neurons, where it stabilizes microtubules, structural elements of the cellular cytoskeleton. However, under pathological conditions, Tau dissociates from the microtubules, becomes hyperphosphorylated, and forms fibrillar amyloid aggregates. The exact mechanisms of this aggregation remain poorly understood. The study of Tau aggregation relies on the in vitro production of amyloid fibers. Due to its high solubility associated with its positive charge, fiber formation requires the addition of polyanionic molecules, called cofactors, such as heparin (a polysaccharide), RNA, or lipids. However, uncertainties remain regarding the exact role of these cofactors: do they simply catalyze aggregation, or are they integrated into the fiber structure? If so, what impact does this have on the morphology of the aggregates? Tau's ability to aggregate in the presence of lipids raises questions about its behavior in relation to the different membranes of neurons. Tau’s interaction with plasma membranes has been demonstrated and may play a role in both physiological and pathological processes. Does Tau, in the presence of anionic lipids, compromise membrane integrity? What about non-anionic lipids? To address these questions, this thesis project combines several biophysical approaches: attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR), atomic force microscopy (AFM), transmission electron microscopy (TEM), and plasmon waveguide resonance (PWR). The study is structured around two main axes: (i) characterizing Tau aggregation in the presence of different anionic cofactors (heparin, RNA, phospholipids) and studying their impact on fiber morphology; (ii) assessing the effect of Tau's interaction with lipid membranes of varying compositions on membrane integrity. The results of this thesis provide new insights into the pathogenic mechanisms of Tau and may contribute to a better understanding of tauopathies as well as the development of therapeutic strategies

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Polozov, Ivan V. "Interactions of class A and class L amphipathic helical peptides with model membranes". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/tape16/PQDD_0006/NQ30110.pdf.

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Dannehl, Claudia. "Fragments of the human antimicrobial LL-37 and their interaction with model membranes". Phd thesis, Universität Potsdam, 2013. http://opus.kobv.de/ubp/volltexte/2013/6814/.

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A detailed description of the characteristics of antimicrobial peptides (AMPs) is highly demanded, since the resistance against traditional antibiotics is an emerging problem in medicine. They are part of the innate immune system in every organism, and they are very efficient in the protection against bacteria, viruses, fungi and even cancer cells. Their advantage is that their target is the cell membrane, in contrast to antibiotics which disturb the metabolism of the respective cell type. This allows AMPs to be more active and faster. The lack of an efficient therapy for some cancer types and the evolvement of resistance against existing antitumor agents make AMPs promising in cancer therapy besides being an alternative to traditional antibiotics. The aim of this work was the physical-chemical characterization of two fragments of LL-37, a human antimicrobial peptide from the cathelicidin family. The fragments LL-32 and LL-20 exhibited contrary behavior in biological experiments concerning their activity against bacterial cells, human cells and human cancer cells. LL-32 had even a higher activity than LL-37, while LL-20 had almost no effect. The interaction of the two fragments with model membranes was systematically studied in this work to understand their mode of action. Planar lipid films were mainly applied as model systems in combination with IR-spectroscopy and X-ray scattering methods. Circular Dichroism spectroscopy in bulk systems completed the results. In the first approach, the structure of the peptides was determined in aqueous solution and compared to the structure of the peptides at the air/water interface. In bulk, both peptides are in an unstructured conformation. Adsorbed and confined to at the air-water interface, the peptides differ drastically in their surface activity as well as in the secondary structure. While LL-32 transforms into an α-helix lying flat at the water surface, LL-20 stays partly unstructured. This is in good agreement with the high antimicrobial activity of LL-32. In the second approach, experiments with lipid monolayers as biomimetic models for the cell membrane were performed. It could be shown that the peptides fluidize condensed monolayers of negatively charged DPPG which can be related to the thinning of a bacterial cell membrane. An interaction of the peptides with zwitterionic PCs, as models for mammalian cells, was not clearly observed, even though LL-32 is haemolytic. In the third approach, the lipid monolayers were more adapted to the composition of human erythrocyte membranes by incorporating sphingomyelin (SM) into the PC monolayers. Physical-chemical properties of the lipid films were determined and the influence of the peptides on them was studied. It could be shown that the interaction of the more active LL-32 is strongly increased for heterogeneous lipid films containing both gel and fluid phases, while the interaction of LL-20 with the monolayers was unaffected. The results indicate an interaction of LL-32 with the membrane in a detergent-like way. Additionally, the modelling of the peptide interaction with cancer cells was performed by incorporating some negatively charged lipids into the PC/SM monolayers, but the increased charge had no effect on the interaction of LL-32. It was concluded, that the high anti-cancer activity of the peptide originates from the changed fluidity of cell membrane rather than from the increased surface charge. Furthermore, similarities to the physical-chemical properties of melittin, an AMP from the bee venom, were demonstrated.
Aufgrund der steigenden Resistenzen von Zellstämmen gegen traditionelle Therapeutika sind alternative medizinische Behandlungsmöglichkeiten für bakterielle Infektionen und Krebs stark gefragt. Antimikrobielle Peptide (AMPs) sind Bestandteil der unspezifischen Immunabwehr und kommen in jedem Organismus vor. AMPs lagern sich von außen an die Zellmembran an und zerstören ihre Integrität. Das macht sie effizient und vor allem schnell in der Wirkung gegen Bakterien, Viren, Pilzen und sogar Krebszellen. Das Ziel dieser Arbeit lag in der physikalisch-chemischen Charakterisierung zweier Peptidfragmente die unterschiedliche biologische Aktivität aufweisen. Die Peptide LL-32 und LL-20 waren Teile des humanen LL-37 aus der Kathelizidin-Familie. LL-32 wies eine stärke Aktivität als das Mutterpeptid auf, während LL-20 kaum aktiv gegen die verschiedenen Zelltypen war. In dieser Arbeit wurde die Wechselwirkung der Peptide mit Zellmembranen systematisch anhand von zweidimensionalen Modellmembranen in dieser Arbeit untersucht. Dafür wurden Filmwaagenmessungen mit IR-spektroskopischen und Röntgenstreumethoden gekoppelt. Circulardichroismus-Spektroskopie im Volumen komplementierte die Ergebnisse. In der ersten Näherung wurde die Struktur der Peptide in Lösung mit der Struktur an der Wasser/Luft-Grenzfläche verglichen. In wässriger Lösung sind beide Peptidfragmente unstrukturiert, nehmen jedoch eine α-helikale Sekundärstruktur an, wenn sie an die Wasser/Luft-Grenzfläche adsorbiert sind. Das biologisch unwirksamere LL-20 bleibt dabei teilweise ungeordnet. Das steht im Zusammenhang mit einer geringeren Grenzflächenaktivität des Peptids. In der Zweiten Näherung wurden Versuche mit Lipidmonoschichten als biomimetisches Modell für die Wechselwirkung mit der Zellmembran durchgeführt. Es konnte gezeigt werden, dass sich die Peptide fluidisierend auf negativ geladene Dipalmitylphosphatidylglycerol (DPPG) Monoschichten auswirken, was einer Membranverdünnung an Bakterienzellen entspricht. Eine Interaktion der Peptide mit zwitterionischem Phosphatidylcholin (PC), das als Modell für Säugetierzellen verwendet wurde, konnte nicht klar beobachtet werden, obwohl biologische Experimente das hämolytische Verhalten zumindest von LL-32 zeigten. In der dritten Näherung wurde das Membranmodell näher an die Membran von humanen Erythrozyten angepasst, indem gemischte Monoschichten aus Sphingomyelin (SM) und PC hergestellt wurden. Die physikalisch-chemischen Eigenschaften der Lipidfilme wurden zunächst ausgearbeitet und anschließend der Einfluss der Peptide untersucht. Es konnte anhand verschiedener Versuche gezeigt werden, dass die Wechselwirkung von LL-32 mit der Modellmembran verstärkt ist, wenn eine Koexistenz von fluiden und Gelphasen auftritt. Zusätzlich wurde die Wechselwirkung der Peptide mit der Membran von Krebszellen imitiert, indem ein geringer Anteil negativ geladener Lipide in die Monoschicht eingebaut wurde. Das hatte allerdings keinen nachweislichen Effekt, so dass geschlussfolgert werden konnte, dass die hohe Aktivität von LL-32 gegen Krebszellen ihren Grund in der veränderten Fluidität der Membran hat und nicht in der veränderten Oberflächenladung. Darüber hinaus wurden Ähnlichkeiten zu Melittin, einem AMP aus dem Bienengift, dargelegt. Die Ergebnisse dieser Arbeit sprechen für einen Detergenzien-artigen Wirkmechanismus des Peptids LL-32 an der Zellmembran.

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Lambert, Eléonore. "Apports de la Microscopie à Force Atomique à l’étude de phénomènes dynamiques en biologie et développement instrumental associé". Thesis, Reims, 2018. http://www.theses.fr/2018REIMS014/document.

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Le Laboratoire de Recherche en Nanosciences EA 4682 s’est récemment équipé de la microscopie à force atomique haute-vitesse (HS-AFM) permettant la visualisation en temps réel des dynamiques d’interactions d’un panel infini d’échantillons biologiques à l’échelle nanométrique. De nombreux champ de recherche nécessite la mise au point de techniques permettant à la fois une imagerie dynamique (vidéomicroscopie) mais également de plus en plus une imagerie haute résolution (microscopie champ proche). Ce couplage a été récemment obtenu grâce au développement de la microscopie à force atomique ultra-rapide. La limitation actuelle de ce microscope ultra-rapide, à savoir l’acquisition d’informations en relation uniquement avec la surface de l’objet biologique étudié, crée un rempart à l’obtention de connaissances nouvelles sur les dynamiques sous-jacentes que renferment certains systèmes biomoléculaires. Pour s’affranchir de cette contrainte, nous nous proposons dans ce projet de faire évoluer notre outil de nanocaractérisation en lui ajoutant des fonctionnalités optiques et des fonctionnalités permettant de faire de la spectroscopie de force. La conduite de ce projet se fera selon un travail de développement instrumental scindé en deux grandes étapes : - l’apport d’outils de microscopie optique conventionnels : FRAP – FRET – FLIM – Fluorescence – TIRFM. Nous couplons ainsi la nanocaractérisation hautement résolue spatialement et temporellement avec des informations intrinsèques de nos échantillons. Cette complémentarité apparaît de plus en plus comme fondamentale dans les demandes des biologistes. - la mise au point de protocoles de fonctionnalisation de leviers AFM afin de réaliser de la spectroscopie de force et ainsi obtenir des informations sur les propriétés mécaniques des échantillons biologiques. Ce projet de recherche sera réalisé au Laboratoire de Recherche en Nanosciences EA 4682, Université de Reims Champagne Ardenne sous la direction du Pr. Michael Molinari et du Dr. Maxime Ewald récemment recruté en tant que maître de conférences (sept. 2013) et qui pu démarrer la thématique de la microscopie AFM haute-vitesse au sein de l’équipe. Il s’effectuera en collaboration avec le Pr. T. Ando du Biophysics Lab’ de l’Université de Kanazawa (Japon) pour la partie instrumentation, et avec le Dr. Gabriel Paës pour l’étude des échantillons biologiques. Les objets étudiés lors de cette thèse seront liés au projet ANR Lignoprog qui vient de démarrer au 1er novembre 2014 porté par Dr. Gabriel Paës (INRA UMR FARE, Reims). Dans ce projet, des échantillons biologiques se doivent d’être caractériser en dynamique. Ils concernent la biomasse lignocellulosique (BL), réseau complexe de polymères constituant les parois végétales (PV). La complexité architecturale et chimique de la BL est un frein à sa conversion industrielle. Pour atteindre ce but, non seulement la fraction cellulosique mais aussi les fractions hémicellulosiques et ligneuses doivent être valorisées, sinon les bio-raffineries ne seront pas compétitives. Le principal challenge à relever est celui du coût élevé et de la relative faible efficacité de l’étape de déconstruction enzymatique de la BL. Avec les fonctionnalités d’imagerie développées dans ce projet, nous espérons apporter des éléments de réponses sur la déconstruction enzymatique. Par ailleurs, même si les objets étudiés seront principalement ceux du projet Lignoprog, une validation du dispositif pourra être réalisée en parallèle sur d’autres échantillons biologiques tels que des cellules vivantes seront envisagées : caractérisation, mise en évidence leur réactivité vis-à-vis des divers paramètres physiologiques du milieu (pH, concentration, composition), corrélation de ces résultats avec leurs propriétés mécaniques
Our laboratory recently acquired a high-speed atomic force microscope (HS-AFM) which enables us to visualize in real time a wide range of biological samples and their dynamics of interaction at nanoscale. Several research fields require the development of new techniques in order to get high resolution imaging and dynamic imaging at the same time. This is why HS-AFM was developed. Its current limitation is that the only data it provides are about the surface which means we can’t get access to what occurs beneath. This is limiting the knowledge we could get about the underlying dynamics of some biomolecular system. In order to overcome this issue, we propose to upgrade this nanocharacterization tool by combining optical microscopy and force spectroscopy. This project of instrumental development will be in two major steps: - the adding of conventional optical microscopy : fluorescence, TIRFM, FRAP, FRET, FLIM. The aim is to nanocharacterize sample with highly spatiotemporal data combined in combination with integral data (fundamental to respond to biological issues) - the development of tip functionalization protocols in order to achieve force spectroscopy and get mechanical properties of biological samples This project will take place at the Laboratory of Research in Nanosciences, EA 4682, University of Reims Champagne Ardennes, under the supervision of Pr. Michael Molinari and Dr. Maxime Ewald who started HS-AFM among our team. We will collaborate with Pr. T. Ando from the Biophysics Lab of Kanazawa University (Japan) for the instrumental part and with Dr. Gabriel Paës for the biological samples. The samples used during this thesis will be linked to an ANR project called Lignoprog directed by Dr. Gabriel Paës (INRA, UMR FARE, Reims) and started on the first of November, 2014. In the project, the dynamical aspect of the biological samples is essential. Indeed, lignocellulosic biomass is a complex network of polymers composing plant cell wall. Its architectural and chemical complexity prevents its industrial conversion. In order to be cost-effective, bio refineries need to valorize all the fractions: cellulose, hemicelluloses and lignins. The major challenge is the high cost and low efficiency of the enzymatic hydrolysis of the lignocellulosic biomass. Our aim is to bring some answer to understand better and improve enzymatic hydrolysis thanks to the HS-AFM and the combination of new functionalities. By the way, the disposal might be validated on other biological samples in parallel, such as live cells in order to characterize them, enlighten their reactivity in response to physiological parameters of the medium (pH, concentration, composition) and correlate the results with mechanical properties

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Bechtella, Leïla. "Molecular analysis of the interactions of the cell-penetrating peptide Penetratin and lipid membranes. Contributions of the lipid PIP2, biophysical approaches and benzophenone photoreactivity in model membranes". Thesis, Sorbonne université, 2019. http://www.theses.fr/2019SORUS045.

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Les peptides vecteurs (CPP) peuvent entrer dans les cellules et y transporter des molécules biologiquement actives. De précédents travaux ont montré que les CPP pouvaient remodeler le cytosquelette d'actine, interagissaient fortement avec les lipides chargés négativement et que le PIP2 pourrait jouer un rôle dans l'internalisation de la Pénétratine. Nos expériences en DSC ont montré que la Pénétratine interagit avec les têtes polaires et influence la fluidité de la membrane de vésicules contenant du PIP2. La présence de PIP2 favorise l’interaction Pénétratine-lipide. De plus, l’estimation de l'affinité de liaison par fluorescence du tryptophane a montré que la Pénétratine a une affinité plus élevée pour le PIP2 que pour la PS. Le photomarquage par affinité couplé à la spectrométrie de masse, à l'aide de peptides fonctionnalisés par une benzophénone (Bzp), a permis d’étudier les interactions non covalentes des CPP et des membranes lipidiques à un niveau moléculaire. Le PIP2 s'est avéré être un partenaire d'interaction de la Pénétratine et a été marqué préférentiellement dans les liposomes contenant du PC, de la PS et du PIP2. Nous avons mis en évidence des réactions secondaires très informatives qui peuvent se produire simultanément lors de l'irradiation UV, dans un unique système biologique : un CPP inséré dans une bicouche lipidique. Ce travail montre comment exploiter de manière originale les différentes réactivités de la Bzp dans le contexte d'une membrane lipidique, informant sur l'interaction CPP/lipide au niveau moléculaire comme la profondeur d'insertion ou la fluidité membranaire au voisinage du CPP
Cell-penetrating peptides (CPP) can cross cell membranes and deliver biologically active molecules into cells. Previous work showed that CPPs could remodel the actin cytoskeleton, interacted strongly with negatively charged lipids and PIP2 could play a role in Penetratin internalization. Our DSC experiments showed that Penetratin interacts with polar head groups and impacts the lipid bilayer fluidity of PIP2-containing liposomes. It indicated that presence of PIP2 in liposomes triggers Penetratin-lipid interaction. Moreover, Penetratin binding affinity for PIP2-containing lipid vesicles, estimated by tryptophan fluorescence, pointed out that Penetratin has a higher affinity for PIP2 than for PS. Affinity photocrosslinking coupled to mass spectrometry, using benzophenone (Bzp)-functionalized peptides, was used to study the non-covalent interactions of CPPs and lipid membranes at a molecular level. PIP2 was found to be a good interaction partner for Penetratin and was preferably labelled in liposomes containing PC, PS and PIP2. We revealed highly informative secondary reactions occurring during UV irradiation that can occur concomitantly in a single biological system: a membrane-active peptide inserted within a phospholipid bilayer. This work shows how to exploit in an original way the different reactivities of Bzp in the context of a lipid membrane, giving access to information on the CPP/lipid interaction at a molecular level such as depth of insertion or membrane fluidity in the CPP vicinity

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Veatch, Sarah Louise. "Liquid immiscibility in model bilayer lipid membranes /". Thesis, Connect to this title online; UW restricted, 2004. http://hdl.handle.net/1773/9772.

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Nomura, Daniela Akiko. "Caracterização estrutural de dispersões aquosas de lipídios aniônicos". Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/43/43134/tde-08052018-005348/.

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É conhecido que a força iônica do meio desempenha um papel fundamental na estrutura de vesículas aniônicas de DMPG (dimiristoil fosfatidilglicerol) em dispersões aquosas. A baixa força iônica (~ 6 mM), as dispersões de DMPG exibem várias características anômalas, que foram interpretadas como a abertura de poros na bicamada ao longo da larga região de transição de fase gel-fluida (de ~ 18°C a 30°C). Aqui, revisitamos o sistema de DMPG em tampão a baixa força iônica, mas com dispersões obtidas após a extrusão por filtros de 100 nm, portanto menos polidispersas. Para enfatizar as interações eletrostáticas entre as cabeças polares dos lipídios, que não estarão blindadas pela presença de sais na solução, estudamos dispersões de DMPG em água pura, de modo a monitorar os agregados presentes na dispersão, e suas interações. As dispersões em água foram caracterizadas antes e depois da extrusão. Para tal, utilizamos diversas técnicas experimentais, em diferentes temperaturas: espalhamento de luz estático (SLS) e dinâmico (DLS), calorimetria diferencial de varredura (DSC), Ressonância Paramagnética Eletrônica (RPE) de marcadores de spin incorporados aos agregados, espalhamento de raios-X a altos e baixos ângulos (WAXS e SAXS), e medidas de viscosidade, turbidez, mobilidade eletroforética e condutividade elétrica. Resultados das várias técnicas com dispersões extrusadas de DMPG em tampão mostraram que o comportamento anômalo é observado de forma similar ao de dispersões não extrusadas. Entretanto, o pico de SAXS em muito baixo ângulo é visto de 5 a 45°C, e não apenas na região de transição de fase, portanto não deve ser modelado como a distância entre poros na bicamada lipídica que se abririam nesta região. A distância de repetição relacionada a este pico diminui na região de transição de fase, e com o aumento da concentração lipídica. Medidas de DSC indicaram que, em água, a região de transição de fase da vesícula de DMPG é ainda mais ampla, começando em torno de 10°C, mas ainda terminando em ~ 30oC. No entanto, a alta condutividade elétrica, viscosidade, mobilidade eletroforética, raio efetivo, e a baixa turbidez, vistas apenas na região de transição de fase do DMPG em tampão, são encontradas até altas temperaturas em água, quando a bicamada lipídica já se encontra na fase fluida. Medidas de RPE e WAXS mostraram a transição da membrana de uma fase mais rígida/imóvel/organizada para uma fase mais frouxa/móvel. Dados de espalhamento de luz, RPE e SAXS mostram que, similar ao DMPG em tampão, em água, o DMPG organiza-se como vesículas esféricas, unilamelares, mas possivelmente menores e mais carregadas, exibindo fortes interações vesícula-vesícula. Nas medidas de SAXS, o pico de Bragg na região de muito baixo ângulo foi visto em todas as temperaturas (de 5 a 60°C), sendo que a distância de repetição diminui para temperaturas maiores do que 10oC. Os resultados obtidos para dispersões em água, reforçam o comportamento anômalo observado anteriormente para dispersões em tampão em baixa força iônica. De acordo com eles, propomos a existência de vesículas altamente deformadas e ionizadas a partir de uma certa temperatura, T1 para o DMPG em água e Tmon em tampão baixa força iônica, sendo que em água a forte repulsão eletrostática PG--PG- levaria a fortes deformações e interações vesícula-vesícula, em uma ampla extensão de temperaturas.
It is known that the ionic strength plays a fundamental role in the structure of DMPG (dimyristoyl phosphatidylglycerol) anionic vesicles in water medium. At low ionic strength (~ 6 mM), DMPG dispersions display several anomalous characteristics, which were interpreted as the opening of bilayer pores along the wide bilayer gel-fluid transition region (from ~ 18°C to 30°C). Here, we revisit DMPG in buffer at low ionic strength, but with dispersions obtained after the extrusion by 100 nm filters, thus less polydisperse. To emphasize electrostatic interactions between the polar head-groups, which will not be shielded by ions in solution, we studied DMPG dispersions in pure water to monitor the aggregates in the dispersion and their interactions. Water dispersions were characterized before and after extrusion. For such, we used several experimental techniques, at different temperatures: light scattering, both static (SLS) and dynamic (DLS); differential scanning calorimetry (DSC); electron spin resonance (ESR) of spin labels incorporated into the aggregates, Small and Wide Angle X-Ray Scattering (SAXS and WAXS); and viscosity, turbidity, electrophoretic mobility and electrical conductivity measurements. Several techniques with extruded dispersions of DMPG in buffer showed that the anomalous behavior is also observed. However, the SAXS peak at very low angles is seen from 5 to 45°C, and not only in the phase transition region, therefore it should not be modeled as the distance of correlated pores in the lipid bilayer that would open in this region. The repeating distance related to this peak decreases in the phase transition region, and with increasing lipid concentration. DSC indicates that, in water, the bilayer gel-fluid transition is even wider, starting around 10oC but still ending ~ 30oC. However, high electric conductivity, viscosity, electrophoretic mobility, effective radius and low turbidity found only in the gel-fluid transition region for DMPG in buffer, are found at higher temperatures in water, when lipid bilayers are already in the fluid state. ESR and WAXS measurements evidenced the transition of the membrane from a more rigid/immobile/organized phase to a more soft/mobile phase. Light scattering, ESR and SAXS data showed that, similar to DMPG in buffer, in water, DMPG is organized as spherical unillamelar vesicles, but possibly smaller, highly charged, displaying strong vesicle-vesicle interactions. With SAXS the Bragg peak at very low angles was seen at all temperatures (from 5 to 60°C) with the repetition distance decreasing at temperatures higher than 10 ° C. The results obtained for water dispersions reinforce the anomalous behavior previously observed for buffer at low ionic strength dispersions. According to them, we propose the existence of highly deformed and ionized vesicles from a certain temperature, T1 for DMPG in water and Tmon in buffer at low ionic strength. In water the strong PG- - PG- electrostatic repulsion would lead to strong deformations and vesicle-vesicle interactions, over a wide range of temperatures.

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De, Ghellinck D'Elseghem Alexis. "Natural and model membranes: structure and interaction with bio-active molecules via neutron reflection". Doctoral thesis, Universite Libre de Bruxelles, 2013. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/209550.

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Dans cette thèse de doctorat, la structure de membranes naturelles et modèles et leurs interactions avec des molécules biologiquement actives ont été étudiées au moyen de la réflectométrie de neutrons. Les lipides naturels ont été extraits de la levure Pichia pastoris, poussée en milieux deutéré et hydrogéné. L’analyse a montré que la quantité relative de phospholipides n’est pas affectée par le changement en composition isotopique du milieu de croissance. Cependant, les cellules de levures deutérées contiennent principalement des acides gras C18 :1 alors que le degré d’insaturation est plus élevé chez les levures hydrogénés. Diminuer la température du milieu de croissance permet d’augmenter le degré d’insaturation des acides gras chez les levures deutérées. Une analyse qualitative des sphingolipides a été réalisée et un protocole pour séparer les fractions phosphocholines et phosphoethanolamine a été établi.

La structure de bicouches composées des lipides de levures a été étudiée par réflectivité de neutrons. La bicouche composée de lipides deutérés polaires a une épaisseur similaire aux bicouches faites de phosphocholines C18:1 synthétiques. En présence de stérols, la rugosité aux interfaces entre les têtes polaires et les chaînes augmente. La bicouche composée de lipides polaires hydrogénés est plus mince que celle deutérée. Ceci est dû à la composition en acides gras beaucoup plus variée et du plus grand nombre d’insaturations. En présence de stérols, l’épaisseur de la bicouche hydrogénée augmente.

L’interaction de ces bicouches avec l’amphotéricine B (AmB) a été étudiée. L’AmB est un antifongique qui interagit fortement avec les membranes contenant de l’ergostérol et moins fortement avec des membranes contenant du cholestérol. Dans tous les cas, les molécules d’AmB forment une couche épaisse et diluée au dessus de la bicouche lipidique. En présence de stérols, les molécules d’AmB pénètrent dans la bicouche et change sa structure selon la composition en acide gras.

La structure de bicouches lipidiques de plante et leurs interactions avec des intermédiaires de synthèse ont aussi été étudiées par réflectivité de neutrons. Des mélanges ternaires de plantes étaient déposés sur silicium et des mélanges quaternaires sur saphir. L’épaisseur de la bicouche composée de mélange ternaire est de 38 Å, tandis que celle du mélange ternaire est de 28 Å, la différence venant probablement d’un effet de substrat. La présence de diacylglycérol (DAG) a comme conséquence d’augmenter l’aire par lipide, et ainsi de changer la conformation des têtes polaires. L’interaction des bicouches de lipide de plante avec l’acide phosphatidique (PA) dans le but d’observer un flip-flop possible a aussi été étudiée mais le PA a tendance à désorbé les bicouches du substrat et aucun mécanisme de flip flop n’a été détecté.

Finalement, la localisation d’une petite molécule, le resvératrol, dans des bicouches modèles a été étudiée. Le resvératrol est connu pour être responsable du « paradoxe français » qui est une corrélation inverse entre la consommation d’aliment gras et un faible taux de maladie cardiaque. Quand le resvératrol est adsorbé à partir de la phase liquide, il induit une réorganisation des têtes polaires. Quand il est déposé sur le substrat en présence des lipides, il est présent à l’interface entre les têtes polaires et les chaines.

Doctorat en Sciences
info:eu-repo/semantics/nonPublished

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11

Wessman, Per. "Physico-Chemical Investigations of, and Characterization of Model Membranes for, Lipid-Peptide Interactions". Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-89432.

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Barrett, Matthew. "Structure and dynamics of model lipid membranes". Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät, 2016. http://dx.doi.org/10.18452/17540.

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Das Peptid Amyloid-beta wird seit vielen Jahren mit der Alzheimer''schen Demenz in Verbindung gebracht, aber die Verbindung zwischen dem Peptid und der Herkunft der Symptome bleibt unklar. Eine neue Hypothese besagt, dass Wechselwirkungen von Mono- oder Oligomeren des Amyloid-beta mit neuronalen Zellmembranen zu Veränderungen der Membran-Doppelschichtsruktur führen und Störungen dynamischer Prozesse in den Membranen verursachen können. Mit Methoden der Röntgen- und Neutronenstreuung wurden die Struktur und Dynamik von Modellmembranen und Änderungen durch den Einfluss des Peptids Amyloid-beta auf die Modellmembranen untersucht. Es konnte gezeigt werden, dass Monomere des Peptidfragments Amyloid-beta 22-40 in anionische Lipidmembranen eingebaut werden. Mittels quasielastischer-inkohärenter Neutronenstreuung wurde die Dynamik von Lipidmembran untersucht. Ein Anteil von 1,5 mol % Amyloid-beta 22-40 in einer Lipidmembran bei 30°C verursacht eine Verringerung der Diffusionskoeffizienten sowohl der Schwerpunktbewegung der Lipide im ns-Bereich als auch der Dynamik der Fettsäurereste im ps-Bereich. Andererseits wird in der Gelphase der Lipidmembran bei 15°C ein Anstieg der Diffusionskoeffizienten beider Prozesse beobachtet. Eine Serie von Lipidproben mit unterschiedlichem Cholesteringehalt und eingelagerten Peptiden Amyloid-beta 1-42 und Amyloid-beta 22-40 wurde Mittels Röntgendiffraktion charakterisiert. Für das Peptid Amyloid-beta 22-40 wurden zwei Positionen gefunden, eine auf der Oberfläche der Membran, eine zweite in der Membran eingelagert. Das Peptid Amyloid-beta 1-42 ist teilweise in die Membran eingelagert und ist in einer 40 mol % Cholesteringehaltige Membrane durch eine einzelne Position modelliert. Zusätzlich wird der Entwurf und die Inbetriebnahme der BerILL Feuchtekammer beschrieben.
The peptide amyloid-beta has long been associated with Alzheimer’s disease; however the link between the peptide and the origin of symptoms is poorly understood. An emerging hypothesis is that monomeric and oligomeric forms of the peptide interact with neuronal membranes, resulting in perturbations in the bilayer structure and in the dynamic processes which take place in the bilayer. Using X-ray and neutron scattering techniques, the structure and dynamics of model lipid membranes and the changes which arise in the presence of amyloid-beta peptide fragments have been studied. Monomers of the peptide fragment amyloid-beta 22-40 were found to intercalate into an anionic lipid bilayer. Through quasi-elastic neutron scattering, dynamics of bilayer lipids were observed. The presence of 1.5 mol % of the peptide results in a decrease in the diffusion coefficients for lipid centre of mass motion on the nanosecond time-scale, as well as for the lipid tail dynamics on the picosecond scale at 30°C. On the other hand, in the gel-phase of the lipid, at 15°C, an increase in the diffusion coefficients for both of these processes was observed. A series of samples with various cholesterol content and either the amyloid-beta 22-40 peptide fragment or the amyloid-beta 1-42 full length peptide was characterized using X-ray diffraction. The amyloid-beta 22-40 peptide was found to populate two positions, on the surface and embedded in the bilayer. The amyloid-beta 1-42 peptide embeds itself into the membrane, and is modelled by a single population for high cholesterol levels (40 mol % cholesterol). In addition, the design and commissioning of the BerILL humidity chamber, a sample environment with precise temperature and humidity control compatible with neutron scattering experiments is presented.

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Brown, Aidan. "A physical study of model biological membranes". Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609720.

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Botelho, Ana Vitoria. "Lipid-protein interactions: Photoreceptor membrane model". Diss., The University of Arizona, 2005. http://hdl.handle.net/10150/280765.

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G-protein coupled receptors (GPCRs) are transmembrane proteins capable of recognizing an astonishing variety of biological signals, ranging from photons of light to hormones, odorants, and neurotransmitters involved in key biological signaling processes. The aim of this work is to identify how lipid-protein interactions involving the membrane bilayer ultimately affect such vital biological functions. Here the relationship between the bilayer thickness, hydrophobic mismatch, and protein aggregation are investigated by expanding the framework of membrane-receptor interactions in terms of a new flexible surface model. Previously, we have shown how coupling of the elastic stress-strain due to mismatch of the spontaneous curvature and hydrophobic thickness at the lipid/protein interface can govern the conformational transitions of membrane proteins. This approach has now been extended to include coupling of the lateral organization of the GPCR rhodopsin to the curvature and area stress and strain of the proteolipid membrane. Rhodopsin was labeled with site-specific fluorophores, and a FRET technique was employed to probe protein association in different lipid environments. Moreover, UV-visible spectroscopy was used for thermodynamic characterization of the conformational change of rhodopsin. Lastly, the deformation of the lipids with and without rhodopsin was probed in terms of acyl chain order parameters and relaxation rates by solid-state NMR methods, giving insight into the lipid deformation. The results showed that optimal receptor activation occurs in phosphatidylcholine bilayers of 20-carbon acyl chain length, hence one can say that metarhodopsin II is likely to adopt an elongated shape. Lipids promoting aggregation, or below their gel to liquid crystalline transition temperature all favor formation of metarhodopsin I. The data also showed that association and activation of rhodopsin do not always correlate. In terms of the extended flexible surface model, the stress due to hydrophobic mismatch is coupled via the effective number of lipids surrounding the protein due to the lateral organization of the membrane. The measured changes in rhodopsin-rhodopsin interactions and membrane influences on the conformation of the protein after photoisomerization may be crucial to understanding physiological regulation of the rod disk membranes. They are relevant to understanding the complexity of biomembranes involved in many cellular mechanisms, including signal transduction.

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PERISSINOTTO, FABIO. "Lipid raft formation and lipid-protein interactions in model membranes". Doctoral thesis, Università degli Studi di Trieste, 2018. http://hdl.handle.net/11368/2919798.

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The biological membranes of eukaryotic organisms contain functional, highly dynamic nano-domains called "lipid rafts" (LRs) which are enriched in cholesterol, sphingolipids and GPI-anchor proteins. They are involved in several biological processes which implicate or are mediated by the plasma membrane. Moreover, LRs seem to have a critical role in the onset of some neurodegenerative diseases such as the Alzheimer’s disease (AD), Parkinson’s disease (PD) and Prion protein disorders. In the last two decades, the complexity of studying such domains in living cells has caused a growing interest in the use and design of artificial membrane models, which mimic the structure and composition of biological membranes. In this context, I promoted the formation and investigated the properties of lipid raft domains in artificial lipid bilayers by exploiting Atomic Force Microscopy (AFM). I compared two different fabrication methods for the production of artificial lipid bilayers, the drop-casting and the direct vesicle fusion techniques. I started from one-component lipid membranes and I progressively moved towards more complex models, as binary and ternary lipid compositions, in order to study the main LRs features in relation to specific biological phenomena, such as protein-lipid interactions involved in particular pathological diseases. The direct vesicle fusion method appeared to be the most suitable approach in term of reproducibility, stability and control of lipid composition. I took advantage from this method for carrying out a morphological characterization of raft-like model membranes composed by phosphocoline (DOPC), sphingomyelin (SM) and cholesterol focusing in particular on lipid phase behavior. Membranes exhibited the coexistence of two lipid phases, the fluid phase made by DOPC, and the solid-ordered phase made by SM and cholesterol, the latter resembling raft-like domains. With selected 3-component lipid systems, I then investigated the distribution of GM1 ganglioside, a LR marker, into my system, demonstrating its preferential localization in the nano-domains and highlighting the feasibility and versatility of model membrane technology. For the first time, I studied the binding of synthetic full-length Prion protein (PrPc), carrying a C-terminal membrane anchor (MA), to LRs domains. The conversion of PrPc into the scrapie isoform PrPsc, which displays high propensity to aggregate leading to cytotoxicity, has been reported to take place into LRs and to be influenced by lipid-anchors. I demonstrated with this study the propensity of this protein to specifically target LR domains of my artificial systems, observing an aggregation process occurring even at low protein concentrations. A comparative analysis with PrPc lacking of MA is however required to assess the role of lipid-anchor into the protein distribution and aggregation. Finally, in the last part of my research I focused on the study of the role of iron ions in the interaction between alpha synuclein (αS) and lipid membranes. αS is the central protein of PD and the presence of amyloid αS fibrils is the main pathological hallmark of the disease. By AFM in combination with attenuated total reflectance infrared (ATR-IR) spectroscopy, I compared the structural behavior of the wild-type (wt) and a mutant form of αS (A53T) in presence of Fe2+ ions and the effect of the iron ions on the interaction with my artificial membrane, and specifically with LRs.

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Malcolmson, Richard Joseph. "Physical studies of cholesterol and cholesteryl esters in model membranes". Thesis, Edinburgh Napier University, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.385910.

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Arouri, Ahmad. "Interaction of antimicrobial peptides with model lipid membranes". kostenfrei, 2009. http://nbn-resolving.de/urn:nbn:de:gbv:3:4-540.

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Leidy, Chad. "Thermotropic behavior of lipid domains in model membranes /". For electronic version search Digital dissertations database. Restricted to UC campuses. Access is free to UC campus dissertations, 2002. http://uclibs.org/PID/11984.

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Witkowski, Thomas, Rainer Backofen i Axel Voigt. "The influence of membrane bound proteins on phase separation and coarsening in cell membranes". Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2014. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-139226.

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A theoretical explanation of the existence of lipid rafts in cell membranes remains a topic of lively debate. Large, micrometer sized rafts are readily observed in artificial membranes and can be explained using thermodynamic models for phase separation and coarsening. In live cells such domains are not observed and various models are proposed to describe why the systems do not coarsen. We review these attempts critically and show within a phase field approach that membrane bound proteins have the potential to explain the different behaviour observed in vitro and in vivo. Large scale simulations are performed to compute scaling laws and size distribution functions under the influence of membrane bound proteins and to observe a significant slow down of the domain coarsening at longer times and a breakdown of the self-similarity of the size-distribution function
Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich

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Witkowski, Thomas, Rainer Backofen i Axel Voigt. "The influence of membrane bound proteins on phase separation and coarsening in cell membranes". Royal Society of Chemistry, 2012. https://tud.qucosa.de/id/qucosa%3A27814.

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A theoretical explanation of the existence of lipid rafts in cell membranes remains a topic of lively debate. Large, micrometer sized rafts are readily observed in artificial membranes and can be explained using thermodynamic models for phase separation and coarsening. In live cells such domains are not observed and various models are proposed to describe why the systems do not coarsen. We review these attempts critically and show within a phase field approach that membrane bound proteins have the potential to explain the different behaviour observed in vitro and in vivo. Large scale simulations are performed to compute scaling laws and size distribution functions under the influence of membrane bound proteins and to observe a significant slow down of the domain coarsening at longer times and a breakdown of the self-similarity of the size-distribution function.
Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

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21

Beard, Jason. "A model of integrative feedback and homeostasis in lipid biosynthesis". Thesis, University of Southampton, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.289506.

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Helmers, Michael. "Kinks in a model for two-phase lipid bilayer membranes". Thesis, University of Oxford, 2011. http://ora.ox.ac.uk/objects/uuid:15343985-1b1c-4123-838d-8e157e837db1.

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In the spontaneous curvature model for two-phase lipid bilayer membranes the shape of vesicles is governed by a combination of an elastic bending energy and an interface energy that penalises the size of phase boundaries. Each lipid phase induces a preferred curvature to the membrane surface, and these curvatures as well as phase boundaries may lead to the development of kinks. In a rotationally symmetric setting we introduce a family of energies for smooth surfaces and phase fields for the lipid components and study convergence to a sharp-interface limit, which depends on the choice of the bending parameters of the phase field model. We prove that, if kinks are excluded, our energies $Gamma$-converge to the commonly used sharp-interface spontaneous curvature energy with the additional assumption of $C^1$-regularity across interfaces. For a choice of parameters such that kinks may appear, we obtain a limit that coincides with the $Gamma$-limit on all reasonable membranes and extends the classical model by assigning a bending energy also to kinks. We illustrate the theoretical result by some numerical examples.

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Bingham, Richard John. "A continuum model of the electroporation of bilayer lipid membranes". Thesis, University of Leeds, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.535113.

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Ariöz, Candan. "Exploring the Interplay of Lipids and Membrane Proteins". Doctoral thesis, Stockholms universitet, Institutionen för biokemi och biofysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-102675.

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The interplay between lipids and membrane proteins is known to affect membrane protein topology and thus have significant effect (control) on their functions. In this PhD thesis, the influence of lipids on the membrane protein function was studied using three different membrane protein models. A monotopic membrane protein, monoglucosyldiacylglyecerol synthase (MGS) from Acholeplasma laidlawii is known to induce intracellular vesicles when expressed in Escherichia coli. The mechanism leading to this unusual phenomenon was investigated by various biochemical and biophysical techniques. The results indicated a doubling of lipid synthesis in the cell, which was triggered by the selective binding of MGS to anionic lipids. Multivariate data analysis revealed a good correlation with MGS production. Furthermore, preferential anionic lipid sequestering by MGS was shown to induce a different fatty acid modeling of E. coli membranes. The roles of specific lipid binding and the probable mechanism leading to intracellular vesicle formation were also investigated. As a second model, a MGS homolog from Synechocystis sp. PCC6803 was selected. MgdA is an integral membrane protein with multiple transmembrane helices and a unique membrane topology. The influence of different type of lipids on MgdA activity was tested with different membrane fractions of Synechocystis. Results indicated a very distinct profile compared to Acholeplasma laidlawii MGS. SQDG, an anionic lipid was found to be the species of the membrane that increased the MgdA activity 7-fold whereas two other lipids (PG and PE) had only minor effects on MgdA. Additionally, a working model of MgdA for the biosynthesis and flow of sugar lipids between Synechocystis membranes was proposed. The last model system was another integral membrane protein with a distinct structure but also a different function. The envelope stress sensor, CpxA and its interaction with E. coli membranes were studied. CpxA autophosphorylation activity was found to be positively regulated by phosphatidylethanolamine and negatively by anionic lipids. In contrast, phosphorylation of CpxR by CpxA revealed to be increased with PG but inhibited by CL. Non-bilayer lipids had a negative impact on CpxA phosphotransfer activity. Taken together, these studies provide a better understanding of the significance of the interplay of lipids and model membrane proteins discussed here.

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Tjörnhammar, Richard. "Classical and Quantum Descriptions of Proteins, Lipids and Membranes". Doctoral thesis, KTH, Teoretisk biologisk fysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-151396.

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In this thesis the properties of proteins and membranes are studied by molecular dynamics simulations. The subject is decomposed into parts addressing free energy calculations in proteins, mechanical inclusion models for lipid bilayers, phase transitions and structural correlations in lipid bilayers and atomistic lipid bilayer models. The work is based on results from large scale computer simulations, quantum mechanical and continuum models. Efficient statistical sampling and the coarseness of the models needed to describe the ordered and disordered states are of central concern. Classical free energy calculations of zinc binding, in metalloproteins, require a quantum mechanical correction in order to obtain realistic binding energies. Classical electrostatic polarisation will influence the binding energy in a large region surrounding the ion and produce reasonable equilibrium structures in the bound state, when compared to experimental evidence. The free energy for inserting a protein into a membrane is calculated with continuum theory. The free energy is assumed quadratic in the mismatch and depend on two elastic constants of the membrane. Under these circumstances, the free energy can then be written as a line tension multiplied by the circumference of the membrane inclusion. The inclusion model and coarse grained particle simulations of the membranes show that the thickness profile around the protein will be an exponentially damped oscillation. Coarse-grained particle simulations of model membranes containing mixtures of phospholipid and cholesterol molecules at different conditions were performed. The gel-to-liquid crystalline phase transition is successively weakened with increasing amounts of cholesterol without disappearing even at a concentration of cholesterol as high as 60%. A united atom parameterization of diacyl lipids was constructed. The aim was to construct a new force field that retains and improves the good agreement for the fluid phase and at the same time produces a gel phase at low temperatures, with properties coherent with experimental findings. The global bilayer tilt obtains an azimuthal value of 31◦ and is aligned between lattice vectors in the bilayer plane. It is also shown that the model yield a correct heat of melting as well as heat capacities in the fluid and gel phase of DPPC.

QC 20140919

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Kluzek, Monika. "Lipid membrane alteration under exposure to alpha-cyclodextrins and pH-responsive pseudopeptide polymers". Thesis, Strasbourg, 2017. http://www.theses.fr/2017STRAE045/document.

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Le développement de nanotransporteurs basés sur des lipides, des polymères et des nanoparticules avec des propriétés «sur mesure» pour augmenter l’efficacité de médicaments, fait l’objet de recherches intensives. Toutefois, la physico-chimie subtile des intéractions polymères-lipides and nanoparticules-lipides présente encore de larges domaines mal compris et de nombreuses questions sans réponse. Ce projet de recherche doctoral utilise des techniques de visualisation (Cryo-MET, LSCM), et de caractérisation (ITC, DSC, SAXS, SANS, QCM-D) avancées pour obtenir des informations nouvelles sur les mécanismes d’interaction entre des Cyclodextrines-α d’autre part, des polymères sensibles au pH d’autre part, et des bicouches modèle de DOPC. La forte influence de ces deux composés sur ces systèmes modèle élucide certains aspects relatifs à la toxicité vis-à-vis des membranes biologiques et suggère de nouvelles approches pour des applications pharmaceutiques
The primary goal of nanomedicine is to improve clinical outcomes. To this end, the development of nanocarriers based on lipids, polymers and nanoparticles with tailor-made properties that enhance the in vivo potency of drugs is a subject of intense research. However, the subtle physical-chemistry of the polymer-lipid and nanoparticle-lipid interactions still present many poorly understood fields of investigation as well as unanswered questions. This doctoral research project utilizes state-of-the-art visualization (Cryo-TEM, LSCM) and characterization (ITC, DSC, SAXS, SANS, QCM-D) techniques to gain novel insights into the interaction between α-Cyclodextrins in the first hand, a pH-responsive polymer in the other hand, and model DOPC bilayers. The strong influence of both compounds on these model systems elucidate some aspects regarding biological membrane toxicity and suggests novel strategies for pharmaceutical applications

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Lundquist, Anna. "Nanosized Bilayer Disks as Model Membranes for Interaction Studies". Doctoral thesis, Uppsala universitet, Fysikalisk kemi, 2008. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-8495.

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PEG-lipid stabilized bilayer disks have been found in lipid mixtures containing polyethylene glycol (PEG)-lipids where the combination of a high bending rigidity and low PEG-lipid/lipid miscibility favours disk formation. The disks are planar and circular in shape and their long-term stability is excellent. Theoretical calculations and experimental observations suggest that the micelle forming PEG-lipid are situated at the rim of the aggregate, protecting the hydrophobic lipid chains in the bulk of the aggregate from contact with water. This thesis deals with fundamental aspects concerning the lipid distribution in the disks, as well as with development, optimization, and initial evaluation of the disks as model membranes in partition and interaction studies. Small angle neutron scattering was used to study the partial segregation of components within the bilayer disk. The experiments verified that the PEG-lipids segregate and accumulate at the bilayer disk rim. The proof of component segregation is important from a fundamental point of view and useful, as exemplified in the below-mentioned study of melittin-lipid interaction, when interpreting partition or binding data obtained from studies based on bilayer disks. Today liposomes are often used as model membranes in partition and interaction studies. Using liposomes to predict, e.g., drug partitioning can however have certain drawbacks. In this thesis the disks were proven to be attractive alternatives to liposomes as model membranes in partition studies. The formation of bilayer disks by a technique based on detergent depletion enabled incorporation of a transmembrane protein in the bilayer disks and opened up for the use of disks as model membranes in membrane protein studies. Further, bilayer disks were used in a comparative study focused on the effect of aggregate curvature on the binding of the peptide melittin. Various techniques were used to perform initial evaluations of the bilayer disks as model membranes. Of these, capillary electrophoresis and biosensor-based technology had not been used before in combination with bilayer disks.

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Fournier, Luc. "A lattice model for the rupture kinetics of lipid bilayer membranes". Thesis, University of Ottawa (Canada), 2002. http://hdl.handle.net/10393/6293.

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We have constructed a model for the kinetics of rupture of membranes under tension, applying physical principles relevant to lipid bilayers held together by hydrophobic interactions. The membrane is characterized by the bulk compressibility (for expansion) K and the thickness 2ht of the hydrophobic part of the bilayer. The model is a lattice model which incorporates stress relaxation, and considers the nucleation of pores at constant area, constant temperature, and constant particle number. The particle number is conserved by allowing multiple occupancy of the sites. A value for the rigidity of the phopholipid tails in the Lalpha liquid phase are found for saturated and unsaturated lipids, and long diblock copolymers. An equilibrium "phase diagram" is constructed as a function of temperature and strain with the pores total surface and distribution as the order parameters. With parameters relevant to saturated phosphatidylcholine (PC) lipid membranes, well defined regions of "no pores", "protopores (non-critical pores)", "rupture" are found. The model also reproduces recent results on super-thick membranes, and on membranes in presence of peptides. Free energy curves as a function of total pore surface are presented for various values of tension and temperature, and the fractal dimension of the pore edge is evaluated.

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Brown, Angela Carin Wrenn Steven Parker. "The effect of lipid composition on cholesterol-rich domain size in model membranes /". Philadelphia, Pa. : Drexel University, 2008. http://hdl.handle.net/1860/2809.

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Fuhrer, Andrew B. "The Role of Lipid Domains and Sterol Chemistry in Nanoparticle-Cell Membrane Interactions". Ohio University / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1596569401131742.

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Soares, Alexandre de Aquino. "Formação de dominios lipidicos em membranas modelo : efeito da dibucaina". [s.n.], 2001. http://repositorio.unicamp.br/jspui/handle/REPOSIP/314161.

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Orientador : Eneida de Paula
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-07-28T10:33:19Z (GMT). No. of bitstreams: 1 Soares_AlexandredeAquino_M.pdf: 12161605 bytes, checksum: 67b1ecf73e749cb80408b243d9326e1d (MD5) Previous issue date: 2001
Resumo: Domínios lipídicos são regiões de composição diferente em uma mesma membrana, quer seja biológica ou sintética. A dinâmica destes domínios pode participar de processos bioquímicos e fisiológicos. Anestésicos locais são fármacos que agem sobre proteínas de membranas por um mecanismo ainda não totalmente conhecido, e cuja potência está fortemente correlacionada com sua hidrofobicidade. Oentre estes, a dibucaína (OSC) é um potente anestésico, com alto coeficiente de partição em favor das membranas, capaz de interferir com diversos processos fisiológicos e de reverter alguns efeitos do íon cálcio, um conhecido indutor de domínios lipídicos. Investigamos a interação da OBC com membranas e o efeito deste AL sobre domínios lipídicos naturais e induzidos por cálcio. As técnicas usadas foram Fluorescência Estática, Ressonância Paramagnética Eletrônica e Microscopia de Fluorescência. Verificamos que a oac possui um pKa de 8,4 e que deve localizar-se na altura do glicerol em membranas compostas por OOPC, o que favorece sua competição com o cálcio pelas cabeças polares de lipídios, e que a forma neutra apresenta maior afinidade pela membrana (_ = 1500) que a protonada (_ = 930). A fluorescência intrínseca da molécula foi analisada em água, em membranas e em soluções de diferentes polaridades; observamos diminuição no rendimento quântico de fluorescência da OSC, concomitante a um deslocamento para o azul nos meios mais apoiares. Experimentos de Ressonância Paramagnética Eletrônica com marcadores de spin do tipo nitróxido em alta concentração (exibindo alargamento dos espectros devido à interação de troca de estado de spin) mostraram que a OBC reverte domínios lipídicos induzidos por cálcio e por magnésio, em sistemas ternários de OOPC: POPA: 5-MeSL. Por microscopia de fluorescência (usando sondas do tipo nitro-benzoxadiol amino, NBO) notamos diferenças evidentes no padrão de formação de domínios lipídicos entre os tratamentos controle, com cálcio, com OBC e com ambos os aditivos, tanto em vesículas unilamelares gigantes quanto em eritrócitos e ghosts de eritrócitos marcados. As diferenças entre os tratamentos foram qualitativamente consistentes para os parâmetros de análise das imagens utilizados. Vimos também aqui o antagonismo entre a OSC e o cálcio e, além disso, uma aparente capacidade da OSC em reverter domínios induzidos por proteínas de membrana. Nossos resultados sugerem que a OSC reverte a separação lateral de fases induzida por cálcio ou proteínas de membrana; esta capacidade da OSC - não'observada em outros anestésicos locais - estaria relacionada à localização preferencial da DSC próximo à cabeça polar dos fosfolipídios. Tal fenômeno pode ser relevante nos processos membranares em que a DSC reconhecidamente interfere, mediados pela alteração do padrão dos domínios lipídicos, dentre os quais a fusão celular e a própria anestesia
Abstract: Lipid domains are compositionally different regions within biological or artificial membranes that may play an important role in physiological and biochemical events as well. Local anesthetics, however, acts upon membrane proteins by an still unknown mechanism and exibit a strong correlation between partition to membranes and anesthetic potency. Among this class of pharmaceuticals, dibucaine (OBC) is a potent one, with a high membrane partition, able to alter several physiological processes and revert some calciuminduced effects on artificial membranes. We have investigated the interaction of neutral and protonated forms of OBC with natural and artificial membranes and its effect upon naturally and calcium-induced lipid domains both in biological and artificial membrane systems. Our approach was based on Static Fluorescence, Electronic Paramagnetic Resonance (EPR) and Fluorescence Microscopy. Our experiments reveal a pKa of 8,4 for OBC and that its aromatic ring may be located close to the glycerol backbone of phospholipid membranes. Surprisingly, OBC emission decreased simultaneously to a classical blue shift when membranes are available or by reducing its solution polarity. EPR experiments strongly suggest that OBC reverts Ca+2- or Mg+2-induced lipid domains. The parameter ais increased in ternary systems composed by OOPC: POPA: 5MeSL when OBC is present, which is antagonic to calcium ion. Fluorescence Microscopy experiments revealed distinct patterns among OBC, Ca+2 or OBC plus Ca+2 treatments, in NBO-PA labeled liposomes, ghosts and erythrocytes. Although Kruskal-Wallis test were not able to tell which treatment differences were statistically significant, those differences were consistent through ali parameters analysed. We could observe antagonism between OBC and Ca+2, besides a further ability of OBC to disrupt naturally occurring lipid domains. These data suggest that OBC is able to revert Ca+2_, Mg+2 or protein-induced domains, for its favoured location close to the phospholipid head. Such phenomena may be relevant in lipid domain mediated processes, which may include anesthesia itself
Mestrado
Bioquimica
Mestre em Biologia Funcional e Molecular

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Morandi, Mattia. "Disruption of model membranes' phase behavior upon interaction with hydrophilic/hydrophobic molecules". Thesis, Strasbourg, 2017. http://www.theses.fr/2017STRAE041/document.

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Ce travail concerne l’altération du comportement de phase de membranes lipidiques lors de leur interaction avec des molécules hydrophiles ou hydrophobes. L’utilisation de sondes moléculaires de fluorescence sensibles à leur micro-environnement constitue un aspect majeur de ce travail. Les techniques de spectroscopie de fluorescence et de microscopie confocale ont été mises à profit pour l’étude du comportement de ces sondes, donnant accès au degré de compacité et d’ordre dans les membranes.Nos résultats montrent que le polystyrène, un plastique rencontré de façon commune dans les régions polluées des océans, présente la capacité de modifier le comportement de phase des membranes lipidiques, entrant notamment en compétition avec le cholestérol.Nous avons montré que la présence élevée de sucres, tel que l’on peut le rencontrer dans certaines situations relevant de la bio-préservation, a pour effet de rompre la qualité de compaction des lipides, et nous avons proposé un nouveau modèle thermodynamique pour interpréter nos résultats.Enfin, les effets sur la membrane de l’incorporation d’un polymère amphiphile comportant un cholestérol greffé ont été étudiés, dans le cadre de l’élaboration de nouvelles stratégies thérapeutiques à base de lipides
This work focuses on the alterations of lipid membrane phase behavior upon interaction with hydrophobic and hydophilic molecules. One major aspect of this thesis is the employement of environment sensitive probes to obtain information on the lipid bilayer packing by means of confocal spectral imaging and fluorescence spectroscopy. Our results show that polystyrene, a commonly found plastic in ocean wastes, has the ability to disrupt the lipid bilayer phase behavior and has a competitive interaction with cholesterol. The presence of high concentration of sugars, relevant in the field of biopreservation, has been found to alter the lipid bilayer packing and a new thermodynamics model has been proposed to complement the experimental results. Finally, the effects of an amphiphilic cholesterol-grafted polymer on model membrane was investigated, providing insight into potential new lipid therapeutic strategies

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Harland, Christopher William 1983. "Desiccation resistance and viscoelasticity in model membrane systems". Thesis, University of Oregon, 2010. http://hdl.handle.net/1794/10993.

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xii, 89 p. : ill. (some col.) A print copy of this thesis is available through the UO Libraries. Search the library catalog for the location and call number.
Lipid membranes are a basic structural element of all cells. They provide a framework for the physical organization of the cell, act as a scaffold for numerous proteins, and serve as the host site for countless chemical reactions integral to cell function. Several key problems in membrane biophysics hinge on reliable methods for measuring membrane material properties. Properties such as rigidity, fluidity, charge density, etc., are important factors that govern membrane structure and function. As such, we need controllable, reliable, and quantitative methods of probing membrane material properties. In pursuit of such methods, we completed two related projects that, while distinct, aimed to create and apply quantitative measures of membrane material properties to current problems in biophysics. The first of these two lines of inquiry centered on the pervasive, pathogenic family of mycobacteria that is known to not only cause several diseases but also to survive prolonged periods of dehydration. We developed an experimental model system that mimics the structure of the mycobacterial envelope consisting of an immobile hydrophobic layer supporting a two-dimensionally fluid, glycolipid-rich outer monolayer. With this system, we show that glycolipid containing monolayers, in great contrast to phospholipid monolayers, survive desiccation with no loss of integrity, as assessed by both fluidity and protein binding, revealing a possible cause of mycobacterial persistence. In the second line of inquiry, we developed another general platform for probing membrane material properties that has produced the first reported observations of viscoelasticity in lipid membranes. We utilized recently developed microrheological techniques on freestanding lipid bilayer systems using high speed video particle tracking. The complex shear modulus of the bilayers was extracted at a variety of temperatures that span the liquid-ordered to disordered phase transition of the membranes. At many temperatures measured, the membranes displayed viscoelastic behavior reminiscent of a Maxwell material, namely elastic at high frequencies and viscous at low frequencies. Moreover, the viscoelastic behavior was suppressed at the critical phase transition temperature where the membranes behave as a purely viscous fluid. Surprisingly, the viscoelastic behavior was found in all of several distinct membrane compositions that were examined.
Committee in charge: Dr. Daniel Steck, Chair; Dr. Raghuveer Parthasarathy, Research Advisor; Dr. Darren Johnson; Dr. Heiner Linke; Dr. John Toner

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Troup, Gregory Marshall Wrenn Steven Parker Dr. "Fluorescence investigation of laterally phase-separated cholesterol rich domains in model lipid membranes using the membrane probe 1-myristoyl-2-[12-[(5-dimethylamino-1-naphthalenesulfonyl)amino]dodecanoyl]-sn-Glycero-3-phosphocholine (A) /". Philadelphia, Pa. : Drexel University, 2004. http://dspace.library.drexel.edu/handle/1860/345.

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Wang, Kathleen F. "Characterizing molecular-scale interactions between antimicrobial peptides and model cell membranes". Digital WPI, 2014. https://digitalcommons.wpi.edu/etd-dissertations/153.

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Due to the escalating challenge of antibiotic resistance in bacteria over the past several decades, interest in the identification and development of antibiotic alternatives has intensified. Antimicrobial peptides (AMPs), which serve as part of the innate immune systems of most eukaryotic organisms, are being researched extensively as potential alternatives. However, the mechanism behind their bactericidal capabilities is not well understood. Previous studies have suggested that AMPs may first attach to the cell membranes, leading to pore formation caused by peptide insertion, lipid removal in the form of peptide-lipid aggregates, or a combination of both mechanisms. In addition to the lack of mechanistic knowledge, a significant hurdle in AMP-based drug development is their potential cytotoxicity to mammalian cells. Understanding AMP interactions with eukaryotic model membranes would allow therapeutics to be tailored for preferential action toward specific classes of bacterial membranes. In this study, we developed novel methods of quartz crystal microbalance with dissipation monitoring (QCM-D) data analysis to determine the fundamental mechanism of action between eukaryotic and bacterial membrane mimics and select membrane-active AMPs. A new technique for creating supported membranes composed entirely of anionic lipids was developed to model Gram-positive bacterial membranes. Atomic force microscopy (AFM) imaging was also used to capture the progression of AMP-induced changes in supported lipid membranes over time and to validate our method of QCM-D analysis. QCM-D and AFM were used to investigate the molecular-scale interactions of four peptides, alamethicin, chrysophsin-3, sheep myeloid antimicrobial peptide (SMAP-29) and indolicidin, with a supported zwitterionic membrane, which served as a model for eukaryotic cell membranes. Since established methods of QCM-D analysis were not sufficient to provide information about these interaction mechanisms, we developed a novel method of using QCM-D overtones to probe molecular events occurring within supported lipid membranes. Also, most previous studies that have used AFM imaging to investigate AMP-membrane interactions have been inconclusive due to AFM limitations and poor image quality. We were able to capture high-resolution AFM images that clearly show the progression of AMP-induced defects in the membrane. Each AMP produced a unique QCM-D signature that clearly distinguished their mechanism of action and provided information on peptide addition to and lipid removal from the membrane. Alamethicin, an alpha-helical peptide, predominantly demonstrated a pore formation mechanism. Chrysophsin-3 and SMAP-29, which are also alpha-helical peptides of varied lengths, inserted into the membrane and adsorbed to the membrane surface. Indolicidin, a shorter peptide that forms a folded, boat-shaped structure, was shown to adsorb and partially insert into the membrane. An investigation of rates at which the peptide actions were initiated revealed that the highest initial interaction rate was demonstrated by SMAP-29, the most cationic peptide in this study. The mechanistic variations in peptide action were related to their fundamental structural properties including length, net charge, hydrophobicity, hydrophobic moment, accessible surface area and the probability of alpha-helical secondary structures. Due to the charges associated with anionic lipids, previous studies have not been successful in forming consistent anionic supported lipid membranes, which were required to mimic Gram-positive bacterial membranes. We developed a new protocol for forming anionic supported lipid membranes and supported vesicle films using a vesicle fusion process. Chrysophsin-3 was shown to favor insertion into the anionic lipid bilayer and did not adsorb to the surface as it did with zwitterionic membranes. When introduced to supported anionic vesicle films, chrysophsin-3 caused some vesicles to rupture, likely through lipid membrane disruption. This study demonstrated that molecular-level interactions between antimicrobial peptides and model cell membranes are largely determined by peptide structure, peptide concentration, and membrane lipid composition. Novel techniques for analyzing QCM-D overtone data were also developed, which could enable the extraction of more molecular orientation and interaction dynamics information from other QCM-D studies. A new method of forming supported anionic membranes was also designed, which may be used to further investigate the behavior of bacterial membranes in future studies. Insight into AMP-membrane interactions and development of AMP structure-activity relationships will facilitate the selection and design of more efficient AMPs for use in therapeutics that could impact the lives of millions of people per year who are threatened by antibiotic-resistant organisms.

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Guidi, Henrique Santos. "Modelos estatísticos para a transição ordem - desordem de camadas lipídicas". Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/43/43134/tde-27032013-130925/.

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Lipídios em solução aquosa formam uma variedade de estruturas diferentes que incluem monocamadas de surfactantes na interface água-ar, conhecidas como monocamadas de Langmuir, como também vesículas unilamelares ou plurilamelares no interior da solução. Sob variação de temperatura, estas estruturas apresentam diferentes fases, observadas através de calorimetria ou variação isotérmica de pressão lateral. Entre as fases apresentadas por estas estruturas, as duas mais importantes se diferenciam pela ordem das cadeias lipídicas. Entendemos que do ponto de vista das fases termodinâmicas, simplificado e qualitativo, monocamadas de Langmuir e bicamadas lipídicas constituem o mesmo sistema físico sob vínculos diferentes. Neste trabalho, desenvolvemos um modelo estatístico para o estudo da transição ordem-desordem destes sistemas, que inclui flutuações de densidade, estas ausentes no modelo de Doniach, de 1980, a base para muitos estudos teóricos para transições de fase de sistemas lipídicos. Flutuações de densidade são fundamentais na descrição de vesículas lipídicas carregadas, compostas de surfactante cuja cabeça polar se dissocia em água. O estudo em laboratório das propriedades térmicas e estruturais de membranas artificiais de lipídios carregados _e relativamente recente, e foi desenvolvido em grande parte no Laboratório de Biofísica do IFUSP. Tais membranas apresentam comportamento distinto das membranas neutras, notoriamente influenciado pela concentração de sal na solução. Isto motivou o desenvolvimento de uma segunda versão do modelo, na qual passamos a descrever a cabeça polar do lipídio em termos de um par de cargas opostas, sendo que a camada lipídica foi acoplada ao modelo primitivo restrito na rede, que desempenha o papel da solução salina. O primeiro modelo foi estudado por aproximação de campo médio e por simulações de Monte Carlo, e o segundo modelo foi investigado apenas através de simulações numéricas. O estudo do modelo carregado foi precedido por uma investigação criteriosa das técnicas de simulação de sistemas com interação Coulombiana, resultando no desenvolvimento de uma metodologia adequada a condições de contorno não isotrópicas e com custo computacional reduzido. Os modelos estatísticos propostos por nós levaram a dois resultados importantes. O modelo para camadas lipídicas neutras é, até hoje, o único modelo estatístico que descreve tanto a transição gel-fluido de bicamadas lipídicas, como a transição líquido condensado - líquido expandido\" de monocamadas de Langmuir, além de descrever também a transição líquido expandido gás na interface água-ar. O modelo para camadas lipídicas que se dissociam em água reproduz a variação abrupta na dissociação, concomitante com a transição ordem-desordem, propriedade que permite interpretar estudos experimentais relativos à condutividade das soluções lipídicas correspondentes.
Lipids in aqueous solution form a variety of different structures which include monolayers of surfactants at the water-air interface, known as Langmuir monolayers, as well as unilamellar or plurilamellar vesicles within the solution. Under temperature variation, these structures display different phases, observed through calorimetry or isothermal variation of lateral pressure. Among the phases presented by these structures, the two most important differ in the order of the lipid chains. From the point of view of the thermodynamic phases, our understanding is that Langmuir monolayers and lipid bilayers constitute the same physical system under different constraints. In this work, we develop a statistical model for the order - disorder transition of lipid bilayers which adds density fluctuations to Doniach\'s 1980 model, which has been considered the basis for many theoretical studies for lipid systems phase transitions. Density fluctuations are essential in the description of the properties of charged vesicles in solution, which consist of surfactants whose polar head dissociates in water. The study in the laboratory of thermal and structural properties of artificial charged lipid membranes is relatively new, and was developed largely in the IFUSP Laboratory of Biophysics. Such membranes exhibit distinct behavior if compared to neutral membranes, notoriously influenced by the solution salt concentration. The experimental investigations motivated us to develop a second model, in which we describe the polar headgroups through a pair of opposite charges. The lipid layer is attached to the lattice restricted primitive model, which plays the role of the saline solution. The first model was studied both through a mean-field approximation as well as through Monte Carlo simulations, whereas the second model was investigated only through numerical simulations. The study of the charged model was preceded by a thorough investigation of the simulation techniques for Coulomb interaction systems, leading to the development of a methodology suitable for non isotropic boundary conditions and with reduced computational cost. The statistical models proposed by us led to two important results. To our knowledge, our model for neutral lipid layers is the only statistical model which, aside from describing simultaneously both the gel-fluid transition of lipid bilayers and the condensed liquid - expanded liquid transition of Langmuir monolayers, also describes the gas- expanded liquid transition at the air-water interface. The model for lipid layers that dissociate in water reproduces the abrupt change in dissociation, concomitant with the order-disorder transition, a property that allows us to interpret experimental studies related to conductivity of the corresponding lipid solutions.

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Winger, Theodore Medard. "Synthesis and characterization of supported bioactive phospholipid membranes : model substrates for biosurface engineering". Diss., Georgia Institute of Technology, 1997. http://hdl.handle.net/1853/9504.

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Lad, Mitaben D. "An investigation of protein and antimicrobial peptide interactions with model lipid membranes". Thesis, University of Reading, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.443364.

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Callow, Philip Austin. "Cationic lipid : DNA complexes - their structure and interactions with model cell membranes". Thesis, King's College London (University of London), 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.400591.

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40

Ergun, Seza. "Structural And Functional Investigation Of The Interaction Of Agomelatine With Model Membranes". Master's thesis, METU, 2012. http://etd.lib.metu.edu.tr/upload/12614997/index.pdf.

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Depression is one of the most commonly seen psychiatric diseases in the population in recent years. Treatment of depression is mainly carried out by psychiatric drugs. In the past few years, agomelatine which is released to the market with a trade name, Valdoxane, has been thought to have far less side effects due to its non-addictive nature, not having trouble when the drug is quitted, and also due to its property of binding only to the specific receptor that the drug interacts with. The action mechanism of agomelatine on the membrane structure has not been clarified yet, for instance, no study has been found in the literature about the interaction of agomelatin with the lipids of biological membranes. In this current study, the interaction of agomelatine with the model membranes of dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylgylcerol (DPPG) and sphingomyelin (SM) is examined by Fourier transform infrared spectroscopy (FTIR) and Differential scanning calorimetry (DSC). DSC and FTIR studies show that, agomelatine shifts the phase transition temperature of DPPC and DPPG multilamellar membrane to the lower degrees, however, it shifts the phase transition temperature of SM membrane to the higher degrees. Agomelatine addition increases the lipid order of the DPPC and SM liposome, whereas, it decreases the lipid order of DPPG liposome. Moreover this drug enhances the membrane fluidity among all types of liposome studied. The increase of v lipid order and increase of fluidity at DPPC and SM liposome indicates domain formation upon drug addition (Vest et al., 2004). This was also confirmed by DSC studies. Agomelatine enhances H bonding capacity of all types of liposomes have been studied. However it has different effects on glycerol backbones of the DPPC and DPPG liposomes. At low agomelatine concentrations the increase in the frequency values indicates a decrease in the hydrogen bonding capacity of the glycerol skeleton of DPPC. In contrast, at high concentrations of agomelatine, a decrease in the frequency values was observed as an indicator of the enhancement of the hydrogen bonding capacity. So it enhances H-bonding capacity at gel phase but lowers it at liquid chrystalline phases. A progressive decreases in Tm was observed at DPPG and DPPC liposomes where it increased the Tm at SM. The pretransition peak is abolished and the Tm peak becomes broad, indicating a larger perturbation to the membrane. These observations indicate the possible interaction of agomelatine with the head group as well. The shoulder seen at the thermograms of DPPC and DPPC liposomes at high doses may indicate the lateral phase separation in to drug-rich and drug-poor domains (D&rsquo
Souza et al., 2009). These results may indicate that agomelatine is partially buried in the hydrocarbon core of the bilayer, interacting primarily with the C2-C8 methylene region of the hydrocarbon chains. All these results highlight the fact that agomelatine interacts around the head group in such a manner that it destabilizes the membrane architecture to a large extent.

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41

Elliott, Richard. "Phase separation in mixed bilayers containing saturated and mono-unsaturated lipids with cholesterol as determined from a microscopic model /". Thesis, Connect to this title online; UW restricted, 2005. http://hdl.handle.net/1773/9675.

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Carter, Ramirez Daniel Marcelo. "Fluorescent and Photocaged Lipids to Probe the Ceramide-mediated Reorganization of Biological Membranes". Thèse, Université d'Ottawa / University of Ottawa, 2013. http://hdl.handle.net/10393/23713.

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This thesis describes the development of novel fluorescent and photocaged lipids, and their application as tools to probe the morphological effects of ceramide (Cer)-mediated membrane reorganization in supported lipid bilayers. Cer is a sphingolipid found in eukaryotic cells that plays a key role in regulating biological processes such as apoptosis, cell-to-cell communication, differentiation and some types of pathogenesis. Sphingolipid and cholesterol-rich lipid rafts in the plasma membrane are thought to be the point of origin for many of this lipid second messenger’s effects. Cer is formed in the exoplasmic leaflet of the plasma membrane via the enzymatic hydrolysis of sphingomyelin. The compositional complexity of biological membranes has prompted the adoption of simpler model systems to study the effects of Cer generation. When it is directly incorporated into model membranes, Cer segregates into highly ordered domains with physical properties that are distinct from those of the surrounding fluid environments. However, enzymatic generation of Cer induces complex and dynamic membrane heterogeneity that is difficult to interpret and reconcile with its direct incorporation. Here I describe the synthesis of 4-nitrobenzo-2-oxa-1,3-diazol-7-yl (NBD)-labelled cholesterol (Chol) and Cer analogs, and their use as probes in model membranes exhibiting liquid-disordered (Ld) and liquid-ordered (Lo) phase coexistence. The Chol probes reproduce the modest enrichment of Chol in Lo membrane domains as well as the Cer-induced displacement of cholesterol. One of the NBD Chol probes is used to provide direct visualization of Chol redistribution during enzymatic Cer generation, and assists in identifying new features as Cer-rich regions. The NBD-labelled Cer quantifies membrane order using orientational order parameter measurements derived from polarized total internal reflection fluorescence microscopy (pTIRFM) images. The probe reports on changes in membrane order upon enzymatic generation of Cer, and indicates a significant increase in the molecular order of Ld membrane regions that is consistent with the redistribution of Chol into these areas. The probe also identifies de novo Cer-rich domains as areas of particularly high molecular order. In the final project area, 6-Bromo-7-hydroxycoumarin-4-ylmethyl (Bhc)-caged Cers are shown to release Cer rapidly and efficiently upon irradiation with near-visible UV light. The caged lipids are then incorporated into supported membranes and photolyzed to release Cer with a high degree of spatial and temporal control. Controlled Cer generation is then used to drive protein-ganglioside clustering in lipid bilayers.

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43

Wang, Xiao Yuan. "The effect of vitamin E on the structure and phase behaviour of phospholipid model membranes". Thesis, King's College London (University of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.369793.

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44

Ayoub, Pierre. "Molecular dynamics study of pyrene excimer formation and oxidation in lipid bilayer models". Thesis, Strasbourg, 2015. http://www.theses.fr/2015STRAE038/document.

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Nous proposons une nouvelle approche pour déterminer le coefficient de diffusion dans des membranes lipidiques se basant sur la formation d'excimères. Alors que les autres modèles statistiques considèrent le système comme un ensemble de points sur un réseau, nous utilisons un modèle à gros grain afin d'étudier des bicouches lipidiques simulées à l'aide du champs de force Martini. Nous déterminons le taux de réaction dépendant du temps à partir des probabilités de survie obtenues a posteriori à l'aide des trajectoires numeriques des bicouches symétriques de DOPC (1,2-Dioleoyl-sn-glycero-3-phosphocholine) et POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) simulées à 283 K et 293 K respectivement. Les dynamiques de collision sont obtenues en distinguant virtuellement les molécules simulées. Les sondes fluorescentes sont supposées semblables aux lipides, et par conséquent, ne modifient pas la dynamique. Nous obtenons une expression générale pour la probabilité de survie en combinant approximation des paires indépendantes et propriétés d'échelle, mais aucune hypothèse n'est faite pour le taux de formation d'excimère. En superposant les intensités d'émission de fluorescence normalisées, déterminées numériquement, aux courbes de titrations expérimentales, nous obtenons deux ensembles de résultats pour le coefficient de diffusion latéral, selon que l'association entre feuillets est autorisée ou pas. Nous utilisons un rayon de capture de 0.5 nm, la distance à partir de laquelle les deux sondes réagissent pour former un excimère. En comparant la dynamique Martini aux expériences de fluorescence, il est possible d'estimer le facteur d'accélération
We propose a novel approach to extract the lateral diffusion coefficient in lipid bilayers using excimer formation. In contrast to previous statistical models that modeled the system as points undergoing jumps from site to site on a lattice, we use coarse-grained molecular dynamics to study lipid bilayers simulated using the Martini force field. We derive time dependent reaction rates from survival probabilities obtained a posteriori from numerically generated trajectories of symmetric DOPC (1,2-Dioleoyl-sn-glycero-3-phosphocholine) and POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) bilayers at 283K and 293K respectively. Collision dynamics are determined by virtually relabeling the simulated molecules. The fluorescent probes are assumed to behave like ordinary membrane lipids and therefore the dynamics remain unaffected. We derive a generalized expression for the survival probability combining independent pairs and size scaling assumptions, but no assumption is made regarding the kinetic rate of the excimer formation process. By fitting the numerically determined normalized fluorescence emission intensities to experimental titration curves, we obtain two sets of results for the lateral diffusion coefficients depending whether interleaflet excimer association is allowed or not. We use a capture radius of 0.5 nm, the distance at which the probes react to form excimers. By relating Martini dynamics to real fluorescence experiments, we estimate the numerical Martini acceleration factor. We also study mixtures of oxidized-non oxidized DOPC and POPC bilayers using a hydroperoxidized model of these lipids for different concentrations of the oxidized component (3.1%, 25% and 50%). Using pair correlation functions, we extract structural information on the systems and determine whether the two components are prone to mixing or not. Finally, we calculate the thermodynamic mixing parameters within the framework of the virial expansion

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45

Zhang, Yawen. "Molecular dynamics simulation study of lipid membranes using coarse-grained models". Thesis, Imperial College London, 2014. http://hdl.handle.net/10044/1/38469.

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In this work we use coarse-grained molecular dynamics simulations to investigate how lipid composition affects the phase transition of phospholipid bilayers. We consider a fully hydrated membrane consisting of saturated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and cholesterol or unsaturated 1,2-dioleoyol-sn-glycero-3-phosphocholine (DOPC). We report structural, dynamic changes occurring in the model bilayer mixtures with varying temperature and composition. Firstly we study the effect of cholesterol on the properties of a DPPC bilayer. We have combined the computations of area per lipid, radial distribution function, chain order parameter and Voronoi construction to quantify the phase transitions, and the coarse-grained (CG) model is found to quantitatively reproduce most of the experimental observations. Based on the changes in the structural and dynamic properties, a temperature-composition phase diagram of DPPC/cholesterol is proposed and compared with the experiments. 'Thread-like' cholesterol clusters in the bilayer at high cholesterol concentrations are observed and the origin of this specific lateral organisation is discussed. To explore the role of the CG bead size, a series of simulations varying the cholesterol cross sectional areas were performed. Parameters obtained from simulation of the different cholesterol isomorphs provide important insight into the microscopic degrees of freedom determining the cholesterol arrangement in the bilayer. The results for the modified cholesterols are further discussed in relation to naturally occurring sterols. Finally, the effect of a mono-unsaturated phospholipid (DOPC) on the main melting phase transition is investigated. This analysis is performed by simulating bilayer systems which were constructed by combining a gel phase DPPC bilayer and a fluid phase DOPC bilayer. The visual observations of the bilayers show that the gel and fluid phases coexist within a wide range of temperature and composition. A temperature-composition phase diagram with phase coexistences is proposed using the information extracted from structural and local composition analysis.

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46

Gąsecka, Paulina. "Polarization resolved nonlinear multimodal microscopy in lipids : from model membranes to myelin in tissues". Thesis, Aix-Marseille, 2015. http://www.theses.fr/2015AIXM4377.

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La microscopie non-linéaire résolue en polarisation est un outil puissant pour accéder à des informations structurelles dans les assemblages biomoléculaires. Les interactions non-linéaires entre matière et lumière induisent des processus complexes où des champs électromagnétiques cohérents interagissent avec les dipôles de transitions moléculaires. Le contrôle de la polarisation des champs électromagnétiques excitateurs et l’étude des réponses non-linéaires induites procurent de riches informations sur la distribution angulaire des molécules présentes dans le volume focal de l’objectif du microscope. Dans cette thèse, nous appliquons cette sensibilité à la polarisation à plusieurs modalités de microscopie cohérentes sans marquage (diffusion cohérente Raman anti-Stokes (CARS), diffusion Cohérente stimulée (SRS)) et à la fluorescence à deux photons (2PEF) afin d’obtenir des informations quantitatives sur la forme de la distribution moléculaire et l’orientation des lipides dans les membranes artificielles, ainsi que dans les membranes biologiques telles que la myéline des tissus de la moelle épinière. Avec cette technique, nous adressons une question fondamentale sur le comportement des ensembles lipidiques dans les membranes et sur l’effet d’autres molécules telles que le cholestérol et les marqueurs fluorescents. Nous démontrons que le CARS résolu en polarisation permet d’accéder à de fines informations sur l’organisation des lipides dans les membranes de la myéline, en deçà de la limite de diffraction
Polarization resolved nonlinear microscopy is a powerful tool to image structural information in biomolecular assemblies. Nonlinear interaction between light and matter lead to complex processes where coherent combinations of optical fields couple to assemblies of molecular transition dipoles. Controlling polarized optical fields and monitoring nonlinear induced signals in a medium can nevertheless bring rich information on molecular orientational organization within the focal spot of a microscope objective. In this PhD thesis we apply this polarization sensitivity to different label-free optical coherent techniques (coherent anti-Stokes Raman scattering (CARS), stimulated Raman scattering (SRS)) and to two-photon fluorescence (2PEF) to retrieve quantitative information on the static molecular distribution shape and orientation of lipids in model membranes and biological membranes such as myelin sheaths in spinal cord tissues. With this technique, we address fundamental questions about lipid packing behavior in membranes, and how it can be affected by other molecules such as cholesterol and the insertion of fluorescent lipid probes. We demonstrate that polarization resolved CARS give access to fine details on lipids arrangement in myelin sheaths, at a sub-diffraction scale. In the context of experimental autoimmune encephalomyelitis disease (EAE) we show, that even at the stage of disruption of the myelin envelope during the demyelination process, lipids multilayers reveal strong capability to preserve their macroscopic self-assembly into highly organized structures, with a degree of disorganization occurring only at the molecular scale

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47

Hussein, Wourood. "Nanoparticle-supported lipid bilayers : A novel approach to introduce curvature in model membranes". Thesis, Umeå universitet, Institutionen för integrativ medicinsk biologi (IMB), 2021. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-179003.

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48

Chebukati, Janetricks Nanjala. "Biophysical characterization of bis(monoacylglycero)phosphate (bmp) model lipid membranes using analytical tools". [Gainesville, Fla.] : University of Florida, 2009. http://purl.fcla.edu/fcla/etd/UFE0041239.

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Yamamoto, Akihisa. "Mesoscopic structural dynamics and mechanics of cell membrane models". 京都大学 (Kyoto University), 2015. http://hdl.handle.net/2433/198928.

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Eichner, Adina [Verfasser]. "New insights into the nanoscaled structure of stratum corneum lipid model membranes applying specifically deuterated lipids in neutron diffraction studies4H : [kumulative Dissertation] / Adina Eichner". Halle, 2017. http://d-nb.info/1148425098/34.

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