Bacteroides fragilis is an important, anaerobic commensal of the human gastro-intestinal tract. As a Gram-negative bacterium, B. fragilis produces a large number of outer membrane vesicles (OMV), spherical globules consisting of outer membrane and periplasmic material, which have a range of potential functions and which are known to be able to deliver their cargo to host dendritic cells (DCs). One of the proteins believed to be packaged into the OMV of B. fragilis is BfUbb (encoded by the ubb gene) which shares 63% homology with human ubiquitin. Ubiquitin is a small, common, eukaryotic protein modifier, which is conjugated to target proteins via a series of activating, conjugating and ligating enzymes, and which has known roles in a wide range of eukaryotic cell processes. Due to key differences between the two proteins, BfUbb has the potential to act as a suicide substrate mimic of ubiquitin. BfUbb was therefore assayed for its ability to interact with ubiquitin E2 conjugating enzymes of the ubiquitylation cascade in vitro, and was found to covalently bind the majority of available enzymes in a DTT-sensitive manner. BfUbb showed a preference for three specific E2 enzymes, all of which are involved in the degradation of mitotic check point proteins, suggesting a role for BfUbb in the inhibition of cell cycle progression and, consequently, tumorigenesis. No binding partners of BfUbb were identified outside of the ubiquitylation cascade, however BfUbb was found to form spontaneous multimers in vitro, the biological function of which is unknown. This study also describes the construction of two sets of plasmids. The first set will allow the expression of untagged and fluorescently tagged forms of BfUbb for purification and use in biochemical assays. The second set will allow the expression of his-tagged and fluorescently tagged forms of BfUbb in mammalian cells, so that the effects of BfUbb on the host epithelial cells may be studied. The proteome of the OMV of B. fragilis was solved using LTQ-Orbitrap mass spectrometry. The identified proteins indicated several putative roles for B. fragilis OMV, including nutrient acquisition and protease inhibition. The suitability of techniques used during the isolation and proteomic analysis of OMV in different studies is discussed. BfUbb-carrying B. fragilis OMV were able to inhibit growth of Salmonella enterica Typhimurium, thus indicating a role for BfUbb in the inhibition of competing, pathogenic bacteria in the gastro-intestinal tract. The conclusions of this study are that the putative roles of both BfUbb and the OMV of B. fragilis may promote both survival of the bacterium and the gastro-intestinal health of the host.

Fusion of viral and host membranes is a key step during infection by membrane-enclosed viruses. The fusion pore plays a critical role, and must dilate to release the viral genome. Prior studies of fusion mediated by influenza A hemagglutinin (HA) revealed ~2-5 nm pores that flickered before dilating to >10 nm. The mechanisms involved are unknown. Here we studied HA-mediated fusion pore dynamics using a novel single-pore assay (supported by a novel, robust, single-cell optical assay for fusion between HA-expressing cells and nanodiscs), combined with computational simulations accessing extraordinarily long (ms) timescales. We measured pores between HA-expressing fibroblasts and bilayer nanodiscs. From pore currents we infer pore size with millisecond time resolution. Unlike previous in vitro studies, the use of nanodiscs limited the membrane contact areas and maximum pore sizes, better mimicking the initial phases of virus-endosome fusion. In wild-type (WT) HA-mediated fusion pores, pores flickered about a mean pore size ~1.7 nm. In contrast, fusion pores formed by GPI-anchored HA nucleated at less than half the WT rate; results were consistent with earlier findings that showed that while GPI-HA pores stabilize at larger initial conductances than WT, they were not able to enlarge beyond their initial size. We developed radically coarse-grained, explicit lipid molecular dynamics simulations of the fusion pore reconstituted with post-fusion, trans HA hairpins. With WT HA, fusion pores were small, similar to experiment. Over time hairpins gradually converted from trans to cis. With lipid-anchored HA, the trans → cis transition was much accelerated. Once most hairpins had converted to cis, because apposing membranes were released, the fusion pore was able to dilate to sizes close to protein-free. Additionally, in crowded simulations with HA densities approximating those found in HA clusters, we found that HA aggregation, promoted by TMD-TMD interactions, delayed fusion pore dilation by inhibiting the trans → cis transition. Our results suggest that pore dilation requires the trans → cis transition. We hypothesize that this transition is accelerated in GPI-HA by the more mobile lipid anchor, and may explain the larger observed nascent fusion pores.

L’électrolyse est une technologie qui permet de répondre à deux problématiques cruciales. D’une part, répondre au besoin en stockage d’énergie liée à l’intégration de sources intermittentes sur les réseaux électriques. D’autre part, répondre à la croissance de la demande en hydrogène, liée aux marchés naissants de l’hydrogène énergie. La nature des besoins liés au développement de la technologie d’électrolyse implique des sollicitations intermittentes dont les impacts quant au fonctionnement du système sont encore méconnus. En ce sens, et face aux manques de la littérature quant à la modélisation à l’échelle système de la technologie d’électrolyse PEM, un nouveau modèle est développé. Pour cela, le formalisme de modélisation graphique Bond Graph est utilisé, notamment pour sa capacité à représenter tout type d’échange énergétique de manière unifiée. Le modèle développé permet de représenter l’intégralité d’un système d’électrolyse PEM, ses différents composants et lois de contrôle associées. Il est validé sur la base du comportement dynamique d’une installation semi-industrielle disponible au CEA. Ce modèle est ensuite utilisé pour identifier et comprendre les enjeux liées à une sollicitation intermittente d’un système d’électrolyse PEM d’un point de vue de l’efficacité du système, de sa flexibilité et de sa capacité de suivi de charge, de sa fiabilité, de sa sûreté ou encore de sa durabilité. Différentes modifications de conception sont simulées et évaluées à la lumière de ces différents enjeux. Finalement, le modèle Bond Graph est exploité d’un point de vue de ses propriétés structurelles afin d’analyser les conditions de surveillabilité d’un système d’électrolyse PEM. / PEM Electrolysis is a technology which to enable to face two major challenges : (i) Fulfill the need of energy storage caused by the integration of intermittent energy sources on electricity networks; (ii) Cope with the growing need of carbon free hydrogen caused by the future market applications of hydrogen energy. These particular needs, regarding electrolysis technology development, involve an intermittent operating mode which impacts on the dynamic behavior of the system remain unknown. Modelling is a critical tool to understand these issues and provide a thorough analysis. State of the art of existing modelling works highlighted that only a few models take into account the dynamic of the whole system including Balance of Plant. Therefore a new dynamic and multiphysic model was developed under Bond Graph formalism. This graphical modelling formalism was selected especially thanks to its ability to represent any kind of power exchange in a unified way. The model enables to represent the whole system including balance of plant and associated control laws. It is validated on the dynamic behavior of an experimental device available in CEA. The model is then used in order to identify and understand the issues related to intermittent operation of a PEM electrolysis system. These issues are related to system efficiency, flexibility, reliability, safety and durability. Regarding these issues, some design changes are simulated and assessed. Finally, the Bond Graph model and its structural properties enable to perform diagnosis and monitorability analyses of a PEM electrolysis system.

No description available.

Tese de doutoramento. Engenharia Química e Biológica. Universidade do Porto. Faculdade de Engenharia. 2011

archives@tulane.edu / N/A / 1 / Savannah Steadman

No description available.

Abstract not available

There is a considerable volume of work available in literature which suggests that the performance of axial-flow hollow fibre membrane modules is limited by poorly distributed flow through the shell-side. This study was commissioned to examine the distribution of shell-side flow and its effect on mass transfer and to compare the performance measured by the axial-flow configuration to that obtained by a commonly used alternative known as the helically-wound module design. Laminar flow and mass transfer models have been developed to examine performance through axial-flow hollow fibre modules. These models also consider deviations from laminar flow in the form of turbulence and hydrodynamically undeveloped flow. Modelling analysis on four fibre bundle cross-sections quantify the extent to which channelling limits flow and mass transfer performance. Experimental flow and mass transfer work with locally fabricated hollow fibre modules demonstrated some inconsistencies with axial laminar flow modelling. Pressure drop and mass transfer results exceeded predictions from modelling. This thesis has hypothesised that fibres in axial-flow hollow fibre modules are not aligned as straight and parallel rods (as assumed in modelling) but interweave. Fibre interweaving results in flows between ducts. Such flows create mixing between ducts which results in more intimate contact between the flow and membrane surface, the consequence being higher pressure drop and higher mass transfer. The implication from this work was that axial flow and mass transfer modelling was limited in its use for characterisation of shell-side performance. The experience with helically-wound hollow fibre membrane modules (also fabricated locally) was that, by deliberately inducing flow between ducts, it was possible to considerably improve mass transfer performance. It was found that, whilst helically-wound modules could not be packed as tightly as axial-flow modules and required more sophisticated fabrication techniques, the benefit in their use arose from a substantial improvement in the level of shell-side mass transfer.

Includes bibliography.