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Leung, Ho-chuen, and 梁浩銓. "A study of H5N1-M2e-based universal influenza vaccine." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2014. http://hdl.handle.net/10722/208568.
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The ectodomain of influenza matrix protein 2 (M2e) may be an ideal candidate in the development of influenza universal vaccine due to its highly conserved property among different subtypes/strains of influenza virus. M2e based vaccines have been extensively studied and potent cross-subtype/strain protections have been reported. However, more and more M2e mutants of influenza virus have been identified in recent years. It is still unclear whether M2e based vaccines are effective against these M2e mutants of influenza virus.
This study first evaluated cross-protection of an M2e tetrameric peptide vaccine based on H5N1 virus strain A/Vietnam/1194/04 (VN/1194-M2e) against lethal challenges of M2e mutants of H5N1 virus strain A/Hong Kong/156/97 (HK/156) and a novel H7N9 virus, because there are 3 or 5 amino acid differences between VN/1194-M2e and HK/156-M2e or VN/1194-M2e and H7N9-M2e. The results showed that the vaccination of VN/1194-M2e did not induce high level of cross-reactive antibodies against HK/156-M2e and just provided poor cross-protection against lethal challenge of HK/156 virus. In contrast, VN/1194-M2e vaccination induced high level of cross-reactive antibodies against H7N9-M2e. Consistently, the vaccination provided good cross-protection against lethal challenge of H7N9 virus. These results strongly suggested that some mutations in M2e, such as mutations at positions 10, 14 and 16 which found in HK/156 M2e, might affect the M2e vaccine efficacy, but some others, such as five mutations found in H7N9-M2e, might not be critical for the M2e immunogenicity.
This study then investigated the relationship between the M2e immunogenicity and amino acid mutations of the M2e. Beside VN/1194-M2e (P0), we synthesized additional 10 M2e mutant peptides which contain different single or multiple mutations. The 3D structures of these M2e peptides were predicted and analyzed. The prediction results showed that group 1 peptides (P0, P10, P14, P16, P18, P20 and P18-20) exhibited either irregular structures or loose hairpin structures which might associate with well exposure of antigenic epitope, whereas group 2 peptides (P10-14, P10-16, P14-16 and P10-14-16) formed tight hairpin structures in which antigenic epitope might bury inside their own secondary structure. Vaccination efficacies of these M2e peptides were evaluated in mice for antibody responses and cross-protection against lethal challenge of VN/1194 and HK/156 viruses. Our results showed that vaccinations of group 1 peptides induced high levels of cross-reactive antibodies against VN/1194-M2e and good cross-protection against lethal challenge of VN/1194 virus. However, vaccinations of group 2 peptides vaccinations induced significantly lower VN/1194-M2e antibody responses and poor cross-protection against lethal challenge of VN/119 virus. Furthermore, both group 1 and group 2 peptides could just induce low levels of cross-reactive antibodies against HK/156-M2e and poor protection against lethal challenge of HK/156 virus.
Although H5N1-M2e tetrameric peptide has been previously shown to protect mice from lethal challenges by different subtypes/strains of influenza virus, this study has shown that certain amino acid variations in M2e could weaken M2e immunogenicity but some others might not. The different secondary structures of M2es may probably associate with their immunogenicity. Our findings have provided valuable information for the development of M2e based universal vaccines.<br>published_or_final_version<br>Microbiology<br>Doctoral<br>Doctor of Philosophy
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Khan, Tila. "Tailored influenza virus vaccines for both the young and old: Vaccine Efficacy of Whole Inactivated Vaccines bearing Immunomodulatory Adjuvants or Multimeric peptides." Diss., Virginia Tech, 2012. http://hdl.handle.net/10919/77130.
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Influenza epidemics and pandemics remain a significant burden to world health and economy. Low efficacy of current inactivated influenza vaccines in the elderly and immunocompromized and the inability to protect against antigenically drifted or shifted strains of influenza virus are the two major problems in influenza vaccine research. To overcome these hurdles, we have utilized an in vitro cell culture vaccine platform, which results in whole inactivated influenza vaccine (WIV) bearing bioactive membrane-anchored immunomodulatory proteins such as cytokines on the virion surface, collectively known as CYT-IVACs (Cytokine bearing-Inactivated Vaccine). In addition, we tested whether a multimeric M2e peptide presented on WIV can serve to enhance immunogenicity and augment protective efficacy of whole virus vaccines. Our panel of cytokines includes IL-2, IL-4, IL-12, IL-23, and Flt3L as well as the multimeric M2e peptide, all fused to the membrane anchoring regions of influenza virus hemagglutinin protein and constitutively expressed in virus permissive MDCK cell line. Subsequent infection with influenza virus results in incorporation of fusion constructs directly into budding progeny virions that are harvested, purified and inactivated to generate distinct CYT-IVAC formulations. Following validation of immunomodulator incorporation, vaccines were tested for in vivo efficacy in either "young adult" or "aged" female Balb/c mice. Our results demonstrate that our CYT-IVAC~IL-12/HA and CYT-IVAC~IL-23/HA serve as potent mucosal adjuvants in young adult mice elicited significantly high levels of mucosal IgA antibodies and afford superior protection against lethal virus challenge. Our Flt3L/HA formulation was the most effective stimulator of systemic anti-viral antibody levels. In "aged" mice a single dose formulation of IL-12 bearing CYTIVAC was superior at affording protection against lethal homotypic virus challenge. Finally, administration of multimeric M2e molecule co-presented on WIV elicited prolonged antibody responses in "young adult mice" and provided cross-protection from challenge with the heterologous influenza A pandemic strain 2009 H1N1. In conclusion, the CYT-IVAC approach represents a novel tailored advancement to current WIV approaches that has the potential to elicit both potent mucosal and systemic immune responses in young and old.<br>Ph. D.
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Elaish, Mohamed Salaheldin Ahmed Nassif Elaish. "Development of universal Influenza vaccine in chicken with insights on the extracellular domain of Matrix protein 2." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1471877010.
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Sneyd, Hannah, and Hannah Sneyd. "Influenza A: Mechanism of Infection and Development of M2 Ion Channel Inhibitors." Thesis, The University of Arizona, 2017. http://hdl.handle.net/10150/626385.
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Influenza viral infection causes several hospitalizations and claims the lives of many people each year. The threat of epidemic and pandemic are more pressing than ever with newly mutated strains developing every year. Understanding the mechanism of infection of influenza can help identify new potential drug targets and help progress the development of antivirals. Currently there are two classes of FDA approved drugs, neuraminidase inhibitors and M2 ion channel inhibitors, to combat influenza infection. Unfortunately, viral resistance to M2 ion channel blockers has caused them to stop being used for treatment. This paper focuses on understanding influenzas ability to mutate and it mechanism of infection to develop new M2 ion channel blockers.
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Duff, Kevin Campbell. "Biophysical studies on influenza A M2 protein." Thesis, University of Edinburgh, 1993. http://hdl.handle.net/1842/19707.
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The influenza A M2 protein and, in particular, the proposed transmembrane domain, has been implicated in viral infectivity at two stges in the replicative cycle: viral uncoating and assembly. An identical function has been proposed for the protein at both points of interest, that is, that M2 acts as a proton channel. In order to work, this hypothesis assumes that M2 possesses a transmembrane domain, presumably in a α-helical conformation, that this domain orientates in a prescribed manner across the bilayer and that, indeed, M2 is able to translocate protons across the aforementioned bilayer. This thesis examines these assumptions experimentally using a synthetic, 25 amino acid peptide representing the proposed transmembrane domain of M2. The work described herein may be divided into three main areas, each employing a separate biophysical technique. Circular dichroism was employed to assign an α-helical secondary structure to the M2 peptide. Neutron diffraction orientated this region precisely in the bilayer. Electrophysiological techniques observed directly, for the first time in viruses, proton translocation. The effects of amantadine, the only drug prescribed for use against influenza A infections, in each of these structural asnd functional investigations has also been recorded, providing revealing insights into the drug's efficacy. M2 has structural analogs in other enveloped viruses and work such as that reported in this thesis may reveal a common pathway of viral infectivity for groups of enveloped viruses, therefore allowing the possibility of broad-spectrum drug therapies.
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Hayhurst, Andrew. "Studies on the influenza A virus M2 protein." Thesis, Imperial College London, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.250090.
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Carpenter, Timothy S. "Simulation studies of the influenza M2 channel protein." Thesis, University of Oxford, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.504314.
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Ma, Huailiang, and 马怀良. "Identification of human annexin A6 as a novel cellular interactant of influenza A virus M2 protein and regulator of virus budding andrelease." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2012. http://hub.hku.hk/bib/B48521747.
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Influenza viruses exploit sophisticated host cell machinery to replicate, causing both seasonal epidemics and unpredictable pandemics. Studying the host cellular factors interacting with conserved domains of viral proteins will help us to identify key host proteins for the virus infection. This will not only strengthen our understanding of the precise mechanisms of the virus life cycle, but also pave new avenues for anti-viral development.
The cytoplasmic tail of M2 ion channel (M2/CT) is one of these highly conserved domains. It is fully accessible to the host cell machinery after fusion of the virus envelope with the endosomal membrane and during the trafficking, assembly, and budding processes. I hypothesized that recruitment of host cellular factors by M2/CT may regulate the M2-dependent stages of the virus life cycle. Through a large scale yeast two-hybrid (Y2H) screen with the M2/CT used as bait, the human annexin A6 was identified as a novel host cell interactant and this interaction was further confirmed by both GST pull-down assay on purified proteins and co-immunoprecipitation assay on virus infected cells.
A functional characterization of this novel interaction demonstrated that depletion of annexin A6 could enhance the virus production, while its overexpression could reduce the virus propagation, which indicates that annexin A6 is a negative regulator of the virus infection. However, I found that the virus infection could not induce any changes of annexin A6 expression. Therefore, the annexin A6-mediated regulation may depend on the subcellular localization where the interaction with M2/CT occurs. To decipher which step of the virus replication is regulated, we dissected the virus life cycle and found that modulation of annexin A6 expression had no effect on the early stages of the virus life cycle or on viral RNA replication but impaired the release of progeny virus, as suggested by delayed or defective budding events observed at the plasma membrane of virus-infected and annexin A6-overexpressing cells during a transmission electron microscopy study. To further decipher the underlying molecular mechanisms, the contribution of annexin A6-mediated plasma membrane lipid rafts reorganization through cholesterol homeostasis modulation and cortical actin cytoskeleton remodeling was also investigated.
In conclusion, here I have identified the human annexin A6 as a novel host cell interactant of M2/CT that negatively modulate the influenza virus infection by impairing the virus budding and release. This work further supports the idea that M2 is a multifunctional protein and is also consistent with the discovery by Rossman et al. that M2/CT mediates the virus budding process (Rossman et al., 2010). This study further emphasizes the importance of host cell interactants of M2/CT in this process. Regarding the biology of annexins, this study also adds a new member of this protein family in the list of regulators of influenza virus infection.<br>published_or_final_version<br>Public Health<br>Doctoral<br>Doctor of Philosophy
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Spearpoint, Philip Anthony. "Structure-function studies of the influenza A M2 proton channel." Thesis, University College London (University of London), 2008. http://discovery.ucl.ac.uk/1446119/.
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The influenza A virus M2 protein is a minor component of the virus membrane, it forms a homotetrameric, pH-activated, proton-selective channel and is the target of the anti-influenza drugs, amantadine and rimantadine. Two projects were undertaken to study the structure-function relationships of M2 and the mechanism of its inhibition. Firstly, an M2 protein with a pH-sensitive GFP fused to the C-terminus was constructed to determine whether it could be used to measure directly M2 proton conduction. The pH probe was responsive to rimantadine-sensitive changes in the pH gradient which were detected but were small. This was in part due to reduced proton conductance of the protein possibly due to altered conformation of the M2 pore, but may also reflect the low sensitivity of the GFP probe to local pH in the vicinity of the plasma membrane. Secondly tryptophan fluorescence studies of purified, E.coli- expressed M2 protein in detergent micelles investigated the mechanism of proton conduction and inhibition by anti-M2 drugs. Shifts in emission maxima and fluorescence intensity measurements relating to the tryptophans in the M2 pore indicated a molecular interaction between histidine 37 (His37) and tryptophan 41 (Trp41). pKa values from these experiments correlated with previously reported proton-mediated activation and permeation of the M2 channel. Acrylamide quenching showed that upon histidine protonation, Trp41 was more solvent accessible while polarisation measurements indicated a more restricted environment for Trp41 at low pH. Red-edge excitation measurements suggested a proteinaceous pore in which water molecules were highly immobile when the channel was closed, yet could freely reorientate upon channel activation. The M2 inhibitor rimantadine reversed these phenomena either fully or partially. Equilibrium and time-resolved measurements of this drug-induced reversal permitted the determination of both association and dissociation rate constants and the affinities of drug interaction with M2. These results provide further support for an allosteric mechanism of amantadine inhibition.
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Rosen, Julia von. "Le transfert culturel comme transformation de discours : Mme de Stae͏̈l interprète de l'esthétique kantienne." Caen, 2003. http://www.theses.fr/2003CAEN1371.
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L'objectif de la présente étude consiste à analyser l'exemple particulièrement intéressant d'un transfert culturel dans le cadre des relations franco-allemandes, à savoir l'interprétation de l'esthétique kantienne que donne Mme de Stae͏̈l dans son oeuvre critique principale, De l'Allemagne. Cette interprétation doit être - telle est la thèse essentielle sur laquelle se fonde la structure et l'approche méthodologique de l'étude - mise en relation avec les positions esthétiques, philosophiques et linguistiques de Mme de Stae͏̈l, afin que tous les aspects essentiels impliqués dans le processus spécifique du transfert culturel puissent être respectés et intégrés dans l'analyse. Le problème fondamental consiste à déterminer les "objets" précis du transfert et la manière dont Mme de Stae͏̈l intègre les éléments de la théorie esthétique de Kant, telle qu'elle est formulée dans la Critique de la faculté de juger, dans son propre discours dans De l'Allemagne. A cet effet, l'analyse est structurée en trois parties :1. Comparaison entre les positions esthétiques de Mme de Stae͏̈l et de Kant ; 2. Analyse des positions épistémologiques et linguistiques de Mme de Stae͏̈l et 3. Analyse détaillée du discours stae͏̈lien sur l'esthétique kantienne. Il s'avère que les positions esthétiques des deux auteurs se distinguent nettement quant à la fonction et au statut qu'ils attribuent à l'esthétique. Dans sa présentation de la théorie kantienne, Mme de Stae͏̈l produit une sorte d'amalgame entre ses propres idées, les idées d'autres auteurs et celles de Kant en les " assimilant ", " romantisant " et " platonisant ". En analysant les stratégies textuelles de Mme de Stae͏̈l dans le contexte de ses positions philosophiques et linguistiques, on peut conclure que l'auteur n'était pas consciente d'avoir modifié aussi fortement les idées du philosophe allemand. Il faut donc constater que le transfert culturel opéré par Mme de Stae͏̈l représente une transformation profonde des idées kantiennes.
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Hansen, R. K. "Biophysical and biochemical investigations of the M2 peptide of influenza A." Thesis, University of Cambridge, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.603662.
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Membrane proteins, especially passive ion channels, are very difficult to characterise using normal biophysical methods. This thesis used the transmembrane domain of the M2 proton channel of Influenza A (M2-TM) as a model system for biophysical investigations. Patch clamping is the most accurate method for assessing ion channel function, but requires rigid liposomes that can withstand mechanical stress, but are not usually optimal for structural investigations. Here a new hydrogen deuterium exchange (HDX) method has been optimised for the analysis of a membrane spanning peptides in lipid vesicles. A simple electrospray ionisation mass spectrometry method is introduced for HDX studies of carefully prepared aqueous proteoliposome samples. 11 backbone amide sites of M2-TM in lipid vesicles were shown to be resistant to HDX for several weeks. By contrast, HDX of M2-TM in methanol or in detergent micelles was complete within an hour, indicating that the peptide adopts a non-nativee conformation. A solution state nuclear magnetic resonance (NMR) strategy relying on quenching the exchange reaction at low pH, lyophilisation and resuspension in acidified methanol was applied to studies of HDX in a transmembrane peptide for the first and revealed site specific information. His16 and Leu17 exchanged much faster than neighbouring residues consistent with a "flip-open" mechanism for proton conduction. Binding of the drug amantadine weakly perturbs the HDX of Ala8 and Ala9 in agreement with neutron scattering data showing an interaction with the N-terminus of the channel. NMR studies investigated details of the binding and release of fluoroamantadine to liposomes in the presence and absence of M2-TM consistent with the drug binding only to the closed state of the channel in DMPC vesicles. HDX studies have great potential to reveal details of the structure, dynamics and intermolecular interactions of membrane proteins.
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BRACCO, GIUSEPPE. "Studio dinamico della mitragliatrice browning m2 hb: ottimizzazione ed influenza dell’affusto." Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2015. http://hdl.handle.net/2108/203094.
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Mandala, Venkata Shiva. "Structure and dynamics of influenza M2 proton channels from solid-state NMR." Thesis, Massachusetts Institute of Technology, 2021. https://hdl.handle.net/1721.1/130804.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, February, 2021<br>Cataloged from the official PDF of thesis.<br>Includes bibliographical references.<br>The A and B strains of influenza virus places a substantial burden on health, causing around 30 million illnesses, several hundred thousand hospitalizations, and a few tens of thousands of deaths every year in the United States alone. Developing novel antiviral and vaccines against influenza requires understanding the proteins employed by these infectious virions. The matrix-2 protein (M2) is an essential viral protein that conducts protons in the endosomes of infected host cells and induces membrane curvature to facilitate virus budding. M2's proton channel activity is encapsulated by a transmembrane domain (TM) that is targeted by the FDA-approved antiviral drugs amantadine and rimantadine to inhibit viral replication. Proton conduction by M2's TM is mediated by a His-xxx-Trp motif conserved in the otherwise disparate M2 from influenza A (AM2) and influenza B (BM2) strains.<br>The histidine selects for protons and activates the channel at low pH, while the tryptophan is responsible for gating and unidirectional conduction from the N-terminus (outside) to the C-terminus (inside). M2 conducts protons across lipid bilayers at a moderate rate of ca. 10-1000 s⁻¹. AM2 is well characterized and several high-resolution structures in a variety of membrane and membrane-mimetic environments are available, yet the mechanism of gating and the rate-limiting step of proton conduction are unknown. A gating-deficient mutant was utilized to determine that asymmetric conduction in AM2 is due to tryptophan blocking activation of histidine from the C-terminal side under low pH. Further, in phospholipid bilayers, AM2 shows two discrete conformations that interconvert on the proton conduction timescale, providing the first experimental evidence for a transporter-like mechanism.<br>In contrast to AM2, BM2 is relatively poorly studied and no high-resolution structures in membranes is available. BM2 shares little sequence homology with AM2, is not inhibited by the antiviral drugs targeting influenza, and conducts protons faster but more bidirectionally than AM2. The first high-resolution structures of membrane-embedded BM2 in the closed and open states are reported. In contrast to the transporter-like motion of AM2, BM2 undergoes a subtler channel-like scissor opening motion, that allows for more efficient proton conduction at the expense of some bidirectional current.<br>by Venkata Shiva Mandala.<br>Ph. D.<br>Ph.D. Massachusetts Institute of Technology, Department of Chemistry
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Zhang, Yang, and 张阳. "Identification of a novel interaction between the M2 protein of influenza A virus and cyclin D3: consequencesfor cell cycle progression." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B46943663.
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Martyna, Agnieszka. "Structure and function of the M2 amphipathic helix in influenza virus membrane scission." Thesis, University of Kent, 2016. https://kar.kent.ac.uk/56663/.
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Influenza A is an enveloped, negative sense RNA virus which causes annual epidemics and major pandemics. Assembly and budding of new viral particles is a complex and multistep process, of which many aspects remain unclear despite many years of research. The Influenza virus M2 protein is a homotetrameric transmembrane protein, containing three domains: ecto domain, transmembrane domain and cytoplasmic tail (CT). In the final stage of budding it has been shown that M2 mediates membrane scission through an amphipathic helix (AH), which is formed by the first 17 amino acids of the protein's CT, however the exact mechanism by which membrane scission is triggered was not known. Using a collection of biochemical and biophysical techniques we have investigated the structure of the M2 AH and assessed its function in viral assembly and budding. We have shown that the M2 AH is formed upon lipid binding and remains unstructured in solution. It preferentially binds to membranes with high positive membrane curvature, which is detected by sensing the associated lipid packing defects. There are many cationic residues in the polar face of the M2 AH which could interact with anionic lipid headgroups, however charged interactions do not significantly affect the M2 AH interactions with the membranes. When inserted into the membrane the M2 AH increases lipid ordering. The M2 AH also induces positive membrane curvature and mediates membrane scission, which is the last step of Influenza virus budding, allowing for release of newly formed virions in to the environment. Membrane scission is a crucial step not only in viral budding but also in many biological processes, such as endocytosis. Many proteins have been associated with cellular membrane scission; however, the underlying molecular details are still not known. We have shown that AHs in some cellular peptides, such as Arf 1, Endophillin A3 and Epsin 1, which mediate membrane scission in biological processes, appear to work in a similar manner to the M2 AH. They represent a new protein motif that is capable of sensing curvature, inducing curvature and altering membrane fluidity, thereby mediating membrane scission and therefore belong to a novel group of AHs.
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Cheung, Kai-wing. "Generation of recombinant influenza A virus without M2 ion channel protein by introducing a point mutation at the 5' end of viral intron." Thesis, Click to view the E-thesis via HKUTO, 2004. http://sunzi.lib.hku.hk/hkuto/record/B31477239.
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Strassburg, Alec N. "Influence of Relative Compaction on Passive Resistance of Abutments with Mechanically Stabilized Earth (MSE) Wingwalls." BYU ScholarsArchive, 2010. https://scholarsarchive.byu.edu/etd/2593.
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Large scale static lateral load tests were completed on a pile cap with wingwalls under several different sand backfill configurations: no backfill, loosely compacted unconfined, loosely compacted slip plane wall confined, loosely compacted MSE wingwall confined, and densely compacted MSE wingwall confined. The relative compaction of the backfill was varied during each test to observe the change in passive resistance provided by the backfill. The wall types were varied to observe the force placed on the walls and the wall displacement as a result of the laterally loaded pile cap and backfill relative compaction. Passive force-displacement curves were generated from each test. It was found that the densely compacted material provided a much greater passive resistance than the loosely compacted material by 43% (251 kips) when confined by MSE walls. The outward displacement of the MSE walls decreased noticeably for the dense MSE test relative to the loose MSE test. Backfill cracking and heave severity also increased as the relative compaction level of the backfill increased. As the maximum passive force was reached, the reinforcement reached their peak pullout resistance. Correlations were developed between the passive pressure acting on the pile cap and the pressure measured on the MSE wingwalls as a function of distance from the pile cap for both loose and dense backfills. The pressure measured on the wingwalls was approximately 3 to 9% of the pressure acting on the pile cap. As the distance from the pile cap increased, the pressure ratio decreased. This result helps predict the capacity of the wingwalls in abutment design and the amount of allowable wall deflection before pullout of the backfill reinforcement occurs. Three methods were used to model the measured passive force-displacement curves of each test. Overall, the computed curves were in good agreement with the measured curves. However, the triaxial soil friction angle needed to be increased to the plane strain friction angle to accurately model both the loose and dense sand MSE and slip plane wall confined tests. The plane strain friction angle was found to be between 9 to 17% greater than the triaxial friction angle.
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Moffat, Jeffrey C. "Properties of conductance and inhibition of proton channel : M2 from influenza A virus and Fo from Escherichia coli ATP synthase /." Diss., CLICK HERE for online access, 2006. http://contentdm.lib.byu.edu/ETD/image/etd1366.pdf.
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Hlabeli, Batloung. "The influence of mobile payments on the choices and functionings of Micro and Small Enterprise (MSE) entrepreneurs in Lesotho." Master's thesis, Faculty of Commerce, 2020. http://hdl.handle.net/11427/32680.
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Background: This study explores the significance of ICT on the capabilities of unbanked MSE entrepreneurs in Lesotho. MSE entrepreneurs are the primary source of income for their families, especially in the context of developing countries. Accordingly, MSEs are fundamental instruments for economic growth, poverty alleviation and source of employment in developing countries. Hence, the country that invests in this business sector is a step closer to accomplishing its MDGs. Purpose: The study explored and empirically assessed the influence of mobile payments among MSE entrepreneurs through a capabilities lens. Given that MSEs are crucial for developing countries, the targeted participants were MSE entrepreneurs from diverse trading industries in Lesotho. Research methodology: The methodology for this study was qualitative. Exploratory and descriptive research methods were used to evaluate the influence of mobile payments on the capabilities of MSE entrepreneurs. This study adopted Kleine's Choice Framework to explore and empirically assess the benefits of mobile payments among MSE entrepreneurs. Data was collected using semi-structured interviews and field notes and analysed using thematic analysis. Key Findings: The findings revealed the use of M-Pesa within the MSE sector enabled microentrepreneurs to accomplish their goals. The findings show that M-Pesa leads to the growth of MSEs. Therefore, entrepreneurs in the MSE business sector are capable of providing food, shelter and clothes for their families through income generated from their enterprises. Additionally, MSE entrepreneurs bank, transfer and receive money through accessible and available mobile payment platforms. However, these entrepreneurs face issues such as network failure, withdrawal amount limits and security risks while using mobile payments. Value of the study: The study contributes to the existing ICT4D and mobile payments literature in Lesotho by discussing the influence of mobile payment through the Choice Framework.
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Almeida, Elaine Aparecida de. "Influencia do contato precoce mãe-filho e do uso da chupeta na prevalencia do aleitamento materno." [s.n.], 2006. http://repositorio.unicamp.br/jspui/handle/REPOSIP/310095.
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Previous issue date: 2006<br>Resumo: Esta pesquisa objetiva estudar a prevalência do aleitamento materno e aleitamento materno exclusivo ao nascer, aos dois, quatro e seis meses de vida dos bebês; testar a associação entre o uso de chupetas e o desmame precoce, e identificar os complementos alimentares predominantes. O estudo é do tipo longitudinal prospectivo, pois os indivíduos foram examinados ao nascer, aos dois, quatro e seis meses. Foram incluídos na pesquisa 199 binômios dos municípios de Mogi Guaçu e Estiva Gerbi, nascidos de parto vaginal ou cesáreo, com peso ao nascer entre 3000g e 3800g, sem anomalias congênitas ou asfixia perinatal, atendidos na Santa Casa de Misericórdia de Mogi Guaçu, onde o contato pele a pele e sucção na primeira hora de vida é rotina. Os dados foram coletados em prontuário médico, entrevistas e visitas domiciliares aos dois, quatro e seis meses. Os dados foram digitados em banco de dados do SPSS e analisados pelo SAS. Foram utilizados os teste de Qui quadrado e de Fisher, considerando significativo p < 0,05. O projeto de pesquisa foi submetido ao Comitê de Ética em Pesquisa da UNICAMP, que apresentou parecer positivo para sua realização. Na conclusão os índices encontrados aos dois meses foram 89,57%; aos quatro meses 91,61%; e aos seis meses 92,85% para o aleitamento materno. Para o aleitamento materno exclusivo foram 52,76%; 37,76% e 22,22% respectivamente. A chupeta se destacou na pesquisa como o fator de maior significância para o desmame precoce, ficando o leite artificial como o alimento introduzido mais precocemente na criança<br>Abstract: This research has as objective studying the prevalence of maternal suckling and exclusive maternal suckling when the baby is two, four and six months old, testing the association between the use of pacifiers and precocious weaning and identify the most used food complements. The study is the prospective longitudinal type, once the individuals were examined when born, then at two, four and six months old. 199 binomials from the cities of Mogi Guaçu and Estiva Gerbi were included in the research, from vaginal or cesarean birth, with birth weights between 3000g and 3800g, without congenital abnormalities or perinatal asphyxia, attended at the Santa Casa de Misericórdia de Mogi Guaçu, where the skin contact and sucking in the first hours of life is routine. The data were collected through medical records, interviews with the mothers and home visits at two, four and six months old. The data were typed at the data bank of the SPSS and analyzed by the SAS. Square Qui and Fisher tests were used, considering as significant the p < 0,05. The research project was sent to the Unicamp Research Ethic Committee which has presented positive opinion for its accomplishment. In the conclusion the indexes found were that at two months old they were 89,57%, at four months old 91,61% and at six months old 92,85% for maternal suckling. For exclusive maternal suckling they were 52,76%, 37,76% and 22,22% respectively. The pacifier was shown as the major significant factor for precocious weaning being the artificial milk the most precocious food introduced for the child<br>Mestrado<br>Saude da Criança e do Adolescente<br>Mestre em Saude da Criança e do Adolescente
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Siche, Stefanie. "Die Proteine HA und M2 von Influenzaviren." Doctoral thesis, Humboldt-Universität zu Berlin, Lebenswissenschaftliche Fakultät, 2016. http://dx.doi.org/10.18452/17499.
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Die Assemblierung von Influenzaviren erfolgt an Rafts der apikalen Wirtszellplasmamembran mit denen das Hämagglutinin (HA) über Acylierungen im C-Terminus und hydrophobe Aminosäuren seiner Transmembrandomäne (TMD) interagiert. M2 besitzt eine cytoplasmatische amphiphile Helix (AH), die ebenso potenzielle Raft-Motive aufweist: Eine Acylierung und Cholesterol-Bindemotive. In dieser Arbeit wurde per Konfokalmikroskopie an polarisierten Zellen, die fluoreszenzmarkierte M2-Varianten exprimierten, gezeigt, dass diese M2-Motive nicht für den apikalen Transport, der vermutlich durch Raft-ähnliche Vesikel erfolgt, benötigt werden. Messungen des Förster-Resonanzenergietransfers über Fluoreszenz-Lebenszeit-Mikroskopie (FLIM-FRET) in der Plasmamembran lebender Zellen, die fluoreszenzmarkiertes HA und M2 koexprimierten, ergaben, dass diese Motive auch nicht für die Interaktion mit den durch HA, in Abhängigkeit von dessen Raft-Motiven, stabilisierten Raft-Domänen notwendig sind. Mittels reverser Genetik konnten infektiöse WSN-Viren mit fehlender Acylierung am Ende der HA-TMD, nicht jedoch Viren ohne die zwei cytoplasmatischen Acylierungen hergestellt werden. Weiterhin ergaben Wachstumsanalysen, dass die Acylierung von HA und M2 für den gleichen Schritt des viralen Replikationszyklus von Bedeutung sind. Für die M2-AH wurde postuliert, dass sie die Membrankrümmung detektiert und durch Insertion in die Wirtszellmembran die Virusabschnürung bewirkt. Infektiöse Viren ohne M2 oder ohne die AH konnten ebenso wie Viren mit M2 mit einer Helix mit reduzierter Amphiphilität in dieser Arbeit nicht hergestellt werden. Allerdings führte die Substitution der AH durch typische krümmungsdetektierende oder modulierende Helices zu Viren, deren Wachstum um zwei bis vier Titerstufen im Vergleich zum Wildtyp reduziert war. Die Helix-Amphiphilität scheint wichtig zu sein, aber auch die Sequenz oder bestimmte Aminosäuren sind offenbar für eine effiziente Virusreplikation notwendig.<br>The assembly of influenza virus particles occurs at the apical plasma membrane of the host cell at membrane rafts which the hemagglutinin (HA) interacts with via acylations in its C-terminal region and via hydrophobic amino acids in the transmembrane domain (TMD). M2 possesses a cytoplasmic amphiphilic helix (AH) that also contains potential raft motifs: an acylation and cholesterol-binding motifs. In this work, confocal microscopy of polarised cells, which were expressing fluorescently labelled M2-variants, demonstrated that these motifs of M2 are not required for apical transport, which is assumed to be mediated by raft-like vesicles. Furthermore, FLIM-FRET (Förster resonance energy transfer measured via fluorescence lifetime imaging microscopy) analyses, performed in the plasma membrane of living cells coexpressing fluorescently labelled HA and M2, revealed that these M2-motifs are not required for association with the large coalesced raft phase organised by HA. In contrast, deleting HA’s raft-targeting features clearly reduced clustering with M2. While the removal of the two cytoplasmic acylations prevented the rescue of infectious virus by reverse genetics, a mutant virus without acylation in the HA-TMD could be rescued. Moreover, growth analyses revealed that the acylations of HA and M2 are important for the same step in the viral replication cycle. It has been postulated that the M2-AH detects membrane curvature and accomplishes membrane scission by inserting into the host cell membrane. Viruses without M2, without the M2-AH or with M2 containing a helix with reduced amphiphilicity could not be produced in this work. However, substituting the AH by typical curvature-sensing or -generating helices led to viruses with two to four orders of magnitude reduced growth as compared to wildtype virus. The amphiphilicity of the helix seems to be important, but also the sequence or specific amino acids appear to be necessary for an efficient virus replication.
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CALZADA, MICHEL. "Influence de la non sphericite sur le comportement spectral d'un aerosol a partir d'une comparaison sphere-spheroide." Toulouse 3, 1989. http://www.theses.fr/1989TOU30026.
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Calcul des proprietes de diffusion des particules spheroidales a l'aide d'une methode approchee hybride entre le modele de rayleigh-gans et le modele de lorentz-mie. Evaluation de l'influence du parametre de forme sur un domaine spectral de 2 a 55 microns
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Liao, Shu-Yu Ph D. Massachusetts Institute of Technology. "Structure and dynamics of full-length M2 protein of influenza A virus from solid-state NMR." Thesis, Massachusetts Institute of Technology, 2017. http://hdl.handle.net/1721.1/113974.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, 2017.<br>Cataloged from PDF version of thesis.<br>Includes bibliographical references.<br>Solid-state nuclear magnetic resonance (SSNMR) has been frequently used to elucidate the structure and dynamics of membrane proteins and fibrils that are difficult to characterize by Xray crystallography or solution NMR. This thesis focuses on the structure determination and the proton conduction mechanism of the full-length matrix protein 2 (M2) of influenza A virus. The M2 membrane protein can be separated into three domains: an N-terminal ectodomain (1-2 1), an cc-helical transmembrane domain (TM) (22-46) connected to an amphipathic helix (AH) and a Cterminal cytoplasmic tail (63-97). The TM domain of M2 is responsible for proton conduction ant the ectodomain has been the target for vaccine development. The cytoplasmic tail has been implicated in M2 interaction with other viral proteins from mutagenesis studies. Given the importance of both N- and C-termini, it is essential to determine the structure and the dynamics of M2FL. Furthermore, we are interested in how the cytoplasmic tail affects proton conduction and the interaction of the anti-viral drug amantadine with M2 in the presence of the C-terminus. Using uniformly ¹³C, ¹⁵N-labeled M2FL, our water-selected 2D ¹³C-¹³C correlation experiment indicated that N- and C- termini are on the surface of the lipid bilayer moreover combining with chemical shift prediction, we determined that these two domains are mostly disordered. Deleting the ectodomain of M2FL (M2(21-97)) proved that a small [beta]-strand is located at the N-terminus only in the DMPC-bound state. The M2 conformation is found to be cholesterol-dependent since [beta]-strand is not found in cholesterol-rich membranes. M2(21-97) shows cationic histidine at higher pH, in contrast to M2TM, indicating that the cytoplasmic tail shifts the His37 pKa equilibria. Quantification of the ¹⁵N intensities revealed two pKa's as opposed to of four in M2TM suggesting cooperative proton binding. A possible explanation is that the large number of positively charged residues in the cytoplasmic tail facilitates proton conduction. The cytoplasmic tail was also found to restore drug-binding as amantadine no longer binds to M2(21-61) a in virus-mimetic membrane. These results have extended our understanding of the influence of the cytoplasmic domain on the structure and proton conduction of M2.<br>by Shu-Yu Liao.<br>Ph. D.
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Scott, Claire. "Structure-guided design of novel inhibitors targeting the drug-resistant M2 proton channel from pandemic 'swine' influenza." Thesis, University of Leeds, 2016. http://etheses.whiterose.ac.uk/13510/.
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Antiviral drugs are essential in the early response to pandemic influenza, whilst effective vaccines are developed. The 2009 pandemic H1N1 (pH1N1) strain, and other currently circulating Influenza A viruses (IAVs), are almost ubiquitously resistant to the licensed antivirals amantadine and rimantadine, which target the M2 proton channel. Amongst the polymorphisms associated with M2 drug resistance, asparagine at position 31 (N31) is the most prevalent. With resistance to neuraminidase-targeted antivirals also on the rise there exists an urgent need to develop new inhibitors targeting resistant strains. Through the generation of a pH1N1 M2 homology in silico model, we have identified the first non-adamantane compounds targeting M2 that are effective against the N31 pH1N1 strain. Controversy exists over the binding site of current adamantane based antivirals to M2, with both lumenal and peripheral sites described in the literature. The novel compounds identified herein using our in silico model were selected based upon a predicted preference for one or other of these binding sites. As such, these compounds represent useful tools to investigate the potential of targeting both sites in combination, which will help to mitigate potential drug resistance. A novel in vitro assay was established to test the functional preferences of these compounds, with the results largely supporting in silico predictions. Combinations of lumenally and peripherally targeted compounds resulted in synergistic effects upon infectious virus titre, whereas this was not the case for combinations of compounds predicted to bind at the same site; in the case of two lumenally targeted compounds, this resulted in antagonistic effects. These data support the presence of two drug target sites within the M2 proton channel complex. Finally, whilst resistance within M2 was readily selected against amantadine analogue M2WJ332, this was not the case for non-adamantane compounds, further emphasising the potential benefits of investigating novel compound classes.
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Andreas, Loren B. "Structure and function of the Influenza membrane protein M2 by magic angle spinning NMR and dynamic nuclear polarization." Thesis, Massachusetts Institute of Technology, 2014. http://hdl.handle.net/1721.1/87466.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, 2014.<br>Cataloged from PDF version of thesis. Vita.<br>Includes bibliographical references.<br>Determination of the 3D structure of membrane proteins is a frontier that is rapidly being explored due to the importance of membrane proteins in regulating cellular processes and because they are the target of many drugs. In addition, measuring and understanding how these proteins interact with ligands and other molecules is of critical importance to the design of the next generation of therapeutic agents. With this motivation, we report new methods for the structural characterization of proteins using magic angle spinning (MAS) nuclear magnetic resonance (NMR) applied to the membrane protein M2 from Influenza A, a small helical transmembrane proton transporter that is the target of adamantane based inhibitors but which is now drug resistant due primarily to the point mutation S31N. The development of techniques that boost the sensitivity of NMR are key to its extension to larger molecules such as membrane proteins, and two such methods, dynamic nuclear polarization (DNP) and proton detection, are applied herein. We report the measurement of the distance between the amine of the inhibitor rimantadine and the pore of M2 using DNP. We have applied recoupling techniques to assign the spectra and measure internuclear distances in the drug resistant S31N mutant of M2 in lipid bilayers. Attenuation of strong proton dipole couplings with 60 kHz spinning has allowed us to detect well-resolved proton spectra, and with the higher receptivity of protons, we measured interhelical distances with a methyl-methyl 4D spectrum. Synthesis of the information resulted in a high resolution structure of S3 IN M2.<br>by Loren B. Andreas.<br>Ph. D.
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Peterson, Emily. "Proteoliposome Proton Flux Assays Establish Net Conductance, pH-Sensitivity, and Functional Integrity of a Novel Truncate of the M2 Ion "Channel" of Influenza A." BYU ScholarsArchive, 2010. https://scholarsarchive.byu.edu/etd/2420.
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A novel truncate of Influenza A M2 protein (residues 22-62), incorporated into a uniquely tailored proteoliposome proton uptake assay, demonstrated proton flux more characteristic of an ion transporter than a traditional ion "channel." The liposome paradigm was essential for testing the conductance activity of this M2 truncate at a range of extraphysiological pHs appropriate for channel vs. transport function determination. In addition to transporter-typical proton flux, M2(22-62) showed the key characteristics of functional integrity: selective proton uptake into liposomes and block of uptake by amantadine. Two sets of proteoliposome proton flux assays were carried out, Set 1 at pH values of 6.5, 6.0. 5.5, 5.0, and 4.5; Set 2 at pH values of 6.25, 6.0, 5.75, 5.5, 5.25, 5.0, and 4.75. Observed flux rates followed a proton transport saturation curve similar to that observed in mouse erythroleukemia cells1. Proton transport was maximal at pH 5.5 in Set 1 (139 H+/second/tetramer) and at pH 5.75 in Set 2 (43 H+/second/tetramer). Amantadine block was strongest at pH 5.5 in Set 1 and 6.25 in Set 2, and apparent desensitization of the protein severely reduced proton flux and amantadine sensitivity below pH 5.5 in both sets of experiments. Decreased external pH increased proton uptake with an apparent pKa of 6 (Set 1) or 6.5 (Set 2). These data indicate acid activation of M2(22-62) between pH 5.5-6, optimal amantadine block between pH 5.5-6.25, and a loss of peptide functionality between pH 5.9-4.7.
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Pouzoulet, Christine. "La construction du modèle de Dante comme poète national de l'Italie romantique : de Mme de Stae͏̈l à Quinet, l'exemple de Claude Fauriel (1772-1844) et du réseau de ses relations." Paris 3, 1996. http://www.theses.fr/1996PA030063.
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Ce travail se propose de montrer comment la rehabilitation de dante a l'epoque romantique et sa sacralisation comme premier poete national de l'italie resultent de la liaison intrinseque de deux nouvelles approches. D'une part, l'emergence d'une mise en perspective historique du fait litteraire chez les ideologues et dans le groupe de coppet, puis le developpement de la nouvelle discipline universitaire des litteratures etrangeres apres 1830, modifient la perception de dante, en le resituant dans son epoque et en eclairant son genie comme creation fondatrice d'une langue et d'une litterature nationales. Dante fait ainsi l'objet d'une recuparation politique et devient un symbole liberal, temoignant des droits de l'italie a vivre comme nation. Ce travail analyse donc corollairement le fonctionnement du modele de dante dans le debat liberal francais sur le destin politique et culturel de l'italie. Cette reflexion comparatiste est menee a l'interieur du reseau des relations de laude fauriel (1772-1844) entre la france, l'italie et l'allemagne. Issu du groupe des ideologues, lie a mme de stael et benjamin constant, puis a victor cousin et a l'ecole des jeunes historiens liberaux de la restauration (guizot et thierry surtout) ainsi qu'a stendhal a son retour de milan en 1821, et a quinet apres 1830, fauriel est le premier titulaire de la chaire de litterature etrangere creee a la sorbonne apres la revolution de 1830. Son cours en 1833-34, dante et les origines de la langue et de la litterature italiennes, manifeste sa double attention aux recherches les plus recentes de la philologie allemande et aux preoccupations esthetiques et politiques des romantiques italiens, avec qui il entretient des liens privilegies par son amitie avec manzoni, son sejour en italie de 1823 a 1825, et ses echanges avec visconti et berchet<br>The present research aims at demonstrating how the rediscovery of dante in the romantic era and his subsequent sacralisation as the national poet of italy resulted from the conjunction of two developments. The emergence of a new historical approach to litterature, as practiced by the ideologues and the coppet group together with the institutionalization in the 1830's of foreign literatures as full-fledged academic disciplines, generated a renewed, historicized conception of dante and enlightened his poetic genius as the founding of a national language and literature. Dante's persona was charged with a new political dimension and made into a symbol of italy's legitimacy in existing as a nation. Therefore this work also consideres the functioning of dante as a model in the literal debate carried out in france on the political and cultural destiny of italy. This comparative approach is worked out through a study of fauriel's intellectual network throughout france, italy, germany. Fauriel was the first professor to hold the chair of foreign literature created at la sorbonne after the 1830 revolution. The course he professed in 1833-1834, dante and the origins of italian language and literature, demonstrates his twofold concern for the most recent research in germany and the aesthetic and political discussions carried by the italian romantics to whom he was cloesely related, through his personal friendship with manzoni, his stay in italy from 1823 to 1825 and his exchanges with visconti and berchet
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Williams, Jonathan K. (Jonathan Kyle). "Solid-state nuclear magnetic resonance investigations of the influenza M2 protein : structure and dynamics characterization of channel gating and conduction." Thesis, Massachusetts Institute of Technology, 2017. http://hdl.handle.net/1721.1/113985.
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Thesis: Ph. D. in Physical Chemistry, Massachusetts Institute of Technology, Department of Chemistry, 2017.<br>Cataloged from PDF version of thesis.<br>Includes bibliographical references.<br>Solid-state nuclear magnetic resonance (ssNMR) spectroscopy is a powerful technique that can be used to probe the structure and dynamics of biomolecules that are intractable to study by other structural biology methods. In this thesis, the mechanisms of conduction and gating are investigated in the influenza AM2 and BM2 proton channels bound to native-like lipid bilayers. Using pH-dependent 15N chemical shifts of the conserved Histidine residue, we have found that the proton conduction mechanism is the same in all of the M2 channels (WT AM2, S3 IN AM2, BM2) studied, and that the conduction cycle involves proton shuttling via hydrogen-bonding with water. For the first time, unambiguous structural restraints and sidechain dynamics were measured for the gating residue Trp4l in the AM2 channel. Trp4l was found to come into close contact with His37, close enough to undergo periodic cation-[pi] interactions that ensure unidirectional proton flow. Water-edited ¹H-¹³C correlation experiments were used to investigate the channel hydration of AM2 and BM2 tetramers, and S12 in the C-terminal BM2 channel was found to have the largest pH-dependent spin-diffusion buildup of all residues studied, indicating that the BM2 serine triplet plays an important role in conduction. New ssNMR methods were developed to probe long-range ¹³C-¹³C distances, and to edit and clean-up crowded spectra of peptides and proteins containing aromatic residues. The ASSET technique removes the aliphatic cross peaks of all residues except for Phe, Trp, Tyr, and His in a two-dimensional ¹³C-¹³C correlation experiment, while a two-dimensional ¹³C-¹³C correlation experiment with gated-decoupling retains only non-protonated 3C resonances. The studies in this thesis have provided insight into the functionally important His and Trp residues in the M2 proton channel of the influenza virus, and provide new avenues for studying other peptides and proteins containing aromatic residues.<br>by Jonathan K. Williams.<br>Ph. D. in Physical Chemistry
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Moffat, Jeffrey C. "Properties of Conductance and Inhibition of Proton Channels: M2 from Influenza A Virus and Fo from Escherichia coli ATP Synthase." BYU ScholarsArchive, 2006. https://scholarsarchive.byu.edu/etd/479.
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Proton channels are essential for many of the processes of life. The influenza A viral protein M2 is responsible for sensing the conditions necessary for viral RNA release. The proton-translocating FoF1 ATPase (ATP synthase) uses a proton gradient to drive adenosine triphosphate (ATP) synthesis. We have directly measured proton uptake in vesicles containing reconstituted M2 or FO by monitoring external pH after addition of valinomycin to vesicles with 100-fold diluted external [K+]. This proton flux assay was utilized to quantify proton flux through single M2 and Fo channels. Contrary to previous reports, proton uptake by M2 was not significantly altered by acidification of the extravesicular pH. We conclude that pH only weakly affects proton flux through M2 in the pH range of 5.4 - 7.0. Theoretical analysis utilized for such vesicle uptake assays illuminates the appropriate time scale of the initial slope and an important limitation that must be placed on inferences about channel ion selectivity. The rise in pH over 10 seconds after ionophore addition yielded time-averaged single channel conductances of 0.35±0.2 aS and 0.72±0.4 aS at pH 5.4 and 7.0 respectively. Such a low time-average conductance implies that M2 is only conductive 10^-6 to 10^-4 of the time. M2 selectivity for hydrogen over potassium is ~10^7. Fo translocates protons across membranes, converting electrochemical energy to rotational inertia. Previous experiments have been partially confounded by a contaminating channel, CL, which co-purifies with Fo and leaks cations. CL activity is shown to not decrease following deletion of the previously uncharacterized yraM open reading frame of E. coli. Fo purified from a deletion strain lacking yraM is just as active as Fo purified from the wild-type strain. Using Fo from the deletion strain, the single-hit hypothesis of DCCD inhibition of passive proton flux through Fo was examined. A DCCD-induced reduction in ATP synthase activity correlates with a reduction in the total initial slope, the number of functional Fo per µg protein, and the single channel proton flux. At least 2 DCCD per Fo are required to totally inactivate passive proton flux. M2 and Fo have similar single channel conductances but different open probabilities.
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Bustamante, Guillermo. "Influence of Pile Shape on Resistance to Lateral Loading." BYU ScholarsArchive, 2014. https://scholarsarchive.byu.edu/etd/5630.
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The lateral resistance of pile foundations has typically been based on the resistance of circular pipe piles. In addition, most instrumented lateral load tests and cases history have involved circular piles. However, piles used in engineering practice may also be non-circular cross-section piles such as square and H piles. Some researchers have theorized that the lateral resistance of square piles will be higher than that of circular piles (Reese and Van Impe, 2001; Briaud et al, 1983; Smith, 1987) for various reasons, but there is not test data to support this claims. To provide basic comparative performance data, lateral load tests were performed on piles with circular, square and H sections. To facilitate comparisons, all the tests piles were approximately 12 inches in width or diameter and were made of steel. The square and circular pipe sections had comparable moments of inertia; however, the H pile was loaded about the weak axis, as is often the case of piles supporting integral abutments, and had a much lower moment of inertia. The granular fill around the pile was compacted to approximately 95% of the standard Proctor maximum density and would be typical of fill for a bridge abutment. Lateral load was applied with a free-head condition at a height of 1 ft above the ground surface. To define the load-deflection response, load was applied incrementally to produce deflection increments of about 0.25 inches up to a maximum deflection of about 3 inches. Although the square and pipe pile sections had nearly the same moment of inertia, the square pile provided lateral resistance that was 20 to 30% higher for a given deflection. The lateral resistance of the H pile was smaller than the other two pile shapes but higher than what it is expected based on the moment of inertia. Back analysis with the computer program LPILE indicates that the pile shape was influencing the lateral resistance. Increasing the effective width to account for the shape effect as suggested by Reese and Van Impe (2001) was insufficient to account for the increased resistance. To provide agreement with the measured response, p-multipliers of 1.2 and 1.35 were required for the square pile and H piles, respectively. The analyses suggest that the increased resistance for the square and H pile sections was a result of increases in both the side shear and normal stress components of resistance. Using the back-calculated p-multipliers provided very good agreement between the measured and computed load-deflection curves and the bending moment versus depth curves.
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Williams, Jonathan K., Alexander A. Shcherbakov, Jun Wang, and Mei Hong. "Protonation equilibria and pore-opening structure of the dual-histidine influenza B virus M2 transmembrane proton channel from solid-state NMR." AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, 2017. http://hdl.handle.net/10150/626055.
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The influenza A and B viruses are the primary cause of seasonal flu epidemics. Common to both viruses is the M2 protein, a homotetrameric transmembrane proton channel that acidifies the virion after endocytosis. Although influenza A M2 (AM2) and B M2 (BM2) are functional analogs, they have little sequence homology, except for a conserved HXXXW motif, which is responsible for proton selectivity and channel gating. Importantly, BM2contains a second titratable histidine, His-27, in the tetrameric transmembrane domain that forms a reverse WXXXH motif with the gating tryptophan. To understand how His-27 affects the proton conduction property of BM2, we have used solid-state NMR to characterize the pH-dependent structure and dynamics of His-27. In cholesterol-containing lipid membranes mimicking the virus envelope, N-15 NMR spectra show that the His-27 tetrad protonates with higher pKa values than His-19, indicating that the solvent-accessible His-27 facilitates proton conduction of the channel by increasing the proton dissociation rates of His-19. AM2is inhibited by the amantadine class of antiviral drugs, whereas BM2 has no known inhibitors. Wemeasured the N-terminal interhelical separation of the BM2 channel using fluorinated Phe-5. The interhelical F-19-F-19 distances show a bimodal distribution of a short distance of 7 angstrom and a long distance of 15-20 angstrom, indicating that the phenylene rings do not block small-molecule entry into the channel pore. These results give insights into the lack of amantadine inhibition of BM2 and reveal structural diversities in this family of viral proton channels.
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Yu, Julie. "Alberto Grau the composer, selected works, and influence upon the Venezuelan and international choral community /." Thesis, connect to online resource, 2007. http://digital.library.unt.edu/permalink/meta-dc-3898.
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Thesis (D.M.A.)--University of North Texas, 2007.<br>System requirements: Adobe Acrobat Reader. Accompanied by 3 recitals, recorded Nov. 17, 2005, Apr. 2, 2006, and May 14, 2006. Includes bibliographical references (p. 51-53).
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ARAUJO, EDVAL G. "Influencia das adicoes de NBC e ligas a base de fosforo na sinterizacao do aco rapido M2." reponame:Repositório Institucional do IPEN, 1993. http://repositorio.ipen.br:8080/xmlui/handle/123456789/10372.
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Slone, Connor. "Influence of composition and processing on the mechanical response of multi-principal element alloys containing Ni, Cr, and Co." The Ohio State University, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=osu1555522223986934.
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Cady, Sarah Diane. "Investigation of the structure and dynamics of the M2 transmembrane peptide of the influenza A virus by solid-state nuclear magnetic resonance." [Ames, Iowa : Iowa State University], 2009. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3389091.
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Savoie, Marie-Pier. "La lettre comme lieu d’invention d’un destin littéraire : le cas de Félicité Angers (Laure Conan)." Master's thesis, Université Laval, 2015. http://hdl.handle.net/20.500.11794/26003.
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Félicité Angers, mieux connue sous son pseudonyme de Laure Conan, est la première Canadienne française à vivre de sa plume. Sa correspondance, d’abord recueillie et annotée par Jean-Noël Dion en 2002 sous le titre J’ai tant de sujets de désespoir, puis complétée par la découverte de douze autres lettres parues en 2007 dans l’édition critique d’Angéline de Montbrun préparée par Nicole Bourbonnais, est constituée aujourd’hui de 362 lettres qui demeurent le seul matériau par lequel nous avons accès à l’intimité de l’écrivaine. Nous pourrons donc saisir son parcours par le biais de cet élan qu’est la pratique de l’écriture épistolaire en sillonnant ses lettres intimes, empreintes d’une sensibilité qui lui est propre, tout autant que ses lettres professionnelles, qui témoignent du processus de son émancipation littéraire. Cette correspondance nous permettra également, en dernière partie, de lever le voile sur les influences féminines de l’écrivaine.
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Menoud, Marie-Anne. "Evolution de l'état physiologique de plants de Rosa hybrida L. , variétés "Mme A. Meilland" et "Beauté", au cours du stockage au froid : influence d'une taille estivale." Clermont-Ferrand 2, 1992. http://www.theses.fr/1992CLF21420.
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L’objectif de ce travail est de mettre en évidence les relations entre les conditions de croissance estivale de plants de rosiers cultivés en pépinière et l'évolution de leur état physiologique au cours d'un stockage en chambre froide après arrachage à l'automne, et d'en évaluer les conséquences sur la reprise après plantation. Des plants de Rosa hybrida L. , variétés Mme A. Meilland et Beauté, ont été soumis ou non à une taille estivale. Cette taille a eu pour effet d'homogénéiser les comportements interindividuels, d'accroitre le nombre de fleurs et de feuilles, ainsi que l'allongement total des rameaux. L'évolution de l'état physiologique (dormance), statut glucidique) de ces plants a été suivie au cours d'un stockage de 16 semaines en chambre froide (température comprise entre 1 et 4c). La dormance des bourgeons est de faible intensité et de courte durée. L'inertie la plus forte a été constatée à l'automne et semble affecter davantage la variété Mme A. Meilland que la variété Beauté, et les plants taillés que les plants témoins. Une durée de stockage de 8 semaines contribue à satisfaire les besoins en froid des bourgeons : les teneurs en amidon du système racinaire décroissent alors que celles des glucides solubles augmentent, suggérant l'existence d'interconversions entre ces glucides. Lorsque le stockage se prolonge au-delà de 8 semaines, les conditions de stockage autorisent la croissance des bourgeons. Dans le même temps, les teneurs en saccharose diminuent et les concentrations en glucose et fructose augmentent. L'évolution du statut glucidique des plants lors du stockage n'a pu être mise en relation avec leur aptitude a reprendre leur croissance après repiquage en conditions contrôlées (12 ou 20c). Les taux de reprise et le pouvoir de régénération racinaire présentent de grandes variations interannuelles (caractéristiques climatiques et édaphiques des sites de culture, date d'arrachage, conditions de culture après le repiquage).
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Mohamed, A. S. Mohamed. "Influence de la valorisation de microfibres végétales sur la formation et la résistance aux cycles de gel-dégel de BAP." Thesis, Cergy-Pontoise, 2011. http://www.theses.fr/2011CERG0550/document.
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L'objectif de ce travail est de mettre en exergue les avantages et les inconvénients d'introduire des microfibres végétales, issues du recyclage du carton, dans les bétons autoplaçants BAP, et plus précisément leur influence sur la durabilité aux cycles gel-dégel. Une méthodologie expérimentale a été mise en place pour la formulation des BAP fibrés devant répondre aux exigences suivantes : classe d'exposition XF2, classe de consistance de type Dmoy=68±2 cm. Elle repose sur la théorie de la compacité maximale pour le dosage des particules solides et sur la méthode du mortier de béton équivalent, MBE, pour le dosage en adjuvants. En partant de la composition du BAP de référence, les microfibres végétales ont été introduites à six pourcentages volumiques distincts par rapport au volume du ciment. Une campagne d'essais expérimentaux réalisée sur les MBE fibrés, a montré que l'introduction des microfibres à un dosage compris entre 21% et 41% améliore leurs propriétés physiques et mécaniques. A l'échelle des BAP, les résultats expérimentaux ont montré que les BAP fibrés aux dosages précédents, BAPF 21% et BAPF 41%, possèdent une porosité et une perméabilité plus faibles que le BAP de référence et par conséquent des caractéristiques mécaniques plus élevées. La résistance au gel-dégel des bétons autoplaçants, BAP de référence, BAPF 21% et BAP formulé avec un entraîneur d'air ainsi que de deux bétons vibrés le premier de référence et second contenant 15% en volume du ciment des microfibres, a été étudiée en soumettant ces bétons à des cycles gel-dégel selon la norme NF P18-425. Les résultats obtenus montrent que les bétons additionnés des microfibres végétales sont plus sensibles aux cycles gel-dégel. L'effet nocif des microfibres s'explique par leur nature hydrophile associée à une faible perméabilité des bétons additionnés. De plus, le rôle des granulats sur la sensibilité à l'action du froid a été discuté. Il a été conclu que les granulats silico-calcaires poreux et fragiles sont à éviter dans les bétons destinés aux environnements gélifs<br>The aim of this work is to emphasize the advantages and disadvantages of introducing vegetable based microfibers, resulting from cardboard recycling, in self compacting concrete SCC, and especially their influence on the frost durability. An experimental methodology has been developed for SCC formulation based on following requirements: class of environmental exposure XF2, slump flow Dmoy = 68 ± 2 cm. It is based also on the maximum packing theory for the determination of solid particles content and the method of concrete equivalent mortar, CEM, for superplasticizer dosage. Starting from the SCC of reference composition, vegetable based microfibers were introduced at six different volumetric percentages related to cement volume. A campaign of experimental tests performed on fibred CEM showed that the introduction of microfibers at a volumetric dosage between 21% and 41% improved physical and mechanical properties. On the SCC scale, the experimental results have shown that fibred SCC at previous dosages, FSCC 21% and FSCC 41%, have porosity and permeability lower than the SCC of reference and consequently higher mechanical properties. Frost resistance of , SCC of reference, FSCC 21% and a SCC formulated with an air-entraining as well as two vibrated concretes, the first of reference and second containing 15% microfibers by volume of cement, was studied by subjecting them to freeze-thaw cycles according to NF P18-425. The results show that microfibers added concretes are more susceptible to frost damage. The harmful effect of vegetable based microfibers is explained by their hydrophilic nature associated to a low permeability of fibred concrete. Furthermore, the role of aggregates on the frost sensitivity was discussed. It was concluded that the porous and fragile lime aggregates should be avoided in concrete intended for cold climate
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Halai, Tatsiana. "Parent company influence on spin-off performance." Master's thesis, [s.n.], 2015. http://hdl.handle.net/10284/4934.
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Dissertação apresentada à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Empresariais<br>Nesta tese de mestrado procurou-se analisar a influência da empresa-mãe no desempenho das spin-off. Definiram-se questões de pesquisa de modo a descobrir as razões da criação de spin-off, para compreender os tipos de relações spin-off e o apoio da empresa “mãe” às spin-off no período de pós-spin, bem como identificar os tipos de recursos e conhecimentos, transferências e fazer avaliação do desempenho das spin-off após s sua criação. A fim de encontrar respostas às questões de pesquisa, realizou-se um estudo exploratória, de design qualitativo, recorrendo-se, poisteriormente, à análise quantitativa e descritiva dos dados. O estudo qualitativo foi desenvolvido com o recurso a entrevistas a sete entrevistados, ocupando cargos administrativos e de gestão em empresas de spin-off de EUA e Canadá.Análise, relizando-se uma análise de conteúdo dos dados. O estudo quantitatico recorreu à análise dos rácios financeiros e relatórios de contas sobre o desempenho spin-off.. Durante a análise dos questionários constatou-se que a maioria dos entrevistados mencionaram a maximização de valor ao acionista como a principal razão da criação de spin-off. Apesar da spin-off ser considerada na literatura como entidade independente, 3 das 7 spin-offs foram consideradas dependentes das suas empresas-mãe em recursos, conhecimento e serviços, enquanto empresa-mãe desempenhava um papel de 'manager', 'fornecedor » e « distribuidor » para spin-off. Maioria dos spin-offs recebeu ajuda da empresa-mãe sob a forma de dinheiro, funcionários e gestão. De entre o conhecimento que foi transferido da empresa-mae para a spin-off, destaca-se o pessoal gestão, técnico, tecnológico, marketing, produção e propriedade intelectual. Com base na análise dos rácios financeiros foi descoberto que a performance da spin-off, que não está relacionada com a empresa mãe, desde o ano de separação, pois continua a gerar perda líquida, apesar de que empresa-mãe de spin- off ainda partilhar com a a spin-off recursos e pessoal. A análise de desempenho com base nos indicadores financeiros da spin-off permitiu concluir que o desempenho spin-off tinha vindo a melhorar ligeiramente desde a separação da empresa-mãe. O serviço financeiro da spin-off tem ambições de se tornar um dos líderes no setor financeiro.<br>In this master thesis was made investigation of parent company’s influence on spin-off performance. Research questions were created in order to find out the reasons of spin-off creation, to define types of spin-off and parent-spin-off relationships in the post-spin-off period, to identify types of resources and knowledge, parent transfers to spin-off and to make evaluation of spin-off in post-spin-off performance. In order to find answer on created research questions qualitative and quantitative methods of analysis were used. Qualitative analysis implemented in the form of open-ended questionnaires was answered by 7 respondents, holding administrative and managerial positions in spin-off companies of USA and Canada. Quantitative analysis was made through financial ratios analysis of financial service and energy spin-off performance. During analysis of questionnaires was found that majority of respondents mentioned shareholder value maximization as the main reason of spin-off creation. Despite the fact, that spin-off considered in the literature like independent entity, 3 of 7 spin-offs were found dependent on their parent companies in resources, knowledge and services, while parent company was in a role of ‘manager’, ‘supplier’ and ‘distributor’ for spin-off. Majority of spin-offs got help from parent in the form of cash, employees and management. Among knowledge that was transferred from parent to spin-off was mentioned managerial, technical, technological, marketing, production and intellectual property. Based on the financial ratios performance analysis of energy spin-off was found out that energy spin-off, unrelated to its parent , since the year of separation continue to generate net loss, despite those fact that parent company of spin-off still shares with it resource and personnel. Based on financial ratio performance analysis of financial service spin-off was found out that spin-off performance had been improving slightly since separation from parent. Financial service spin-off have plans to become one of the leaders in the financial industry.
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Millet, Patrice. "Supraconducteurs à haute température critique (Familles 1 2 3 et 2 2 1 2) : influence de la substitution de terres rares : synthèses, structures (R.X), microstructures (M.E.) et propriétés physiques." Toulouse 3, 1990. http://www.theses.fr/1990TOU30012.
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Ce travail est focalise sur les oxydes supraconducteurs a haute temperature critique. La premiere partie est relative a la synthese d'une phase cocktail (ln)ba#2cu#3o#6#+#x ou (ln) represente un melange equimolaire de cinq elements de terres rares. Des monocristaux ont ete obtenus. Il est montre a l'aide de differentes techniques qu'une bonne distribution statistique des terres rares est realisee indiquant que le melange se comporte comme une pseudo terre rare. L'etude de l'evolution de la temperature critique en fonction du rayon ionique de la terre rare montre que celle-ci peut etre modulee en choisissant un cocktail approprie et qu'il est possible d'obtenir des phases supraconductrices, au-dessus de la temperature de l'azote liquide, contenant des taux eleves de lanthane. La seconde partie est consacree a la substitution d'elements de terres rares au calcium dans la phase bi#2sr#2cacu#2o#8#+#x. Il est montre que le lanthane se substitue preferentiellement dans le site du strontium alors que l'erbium et l'ytterbium occupent le site du calcium. L'etude de la variation de t#c en fonction du taux de substitution y, dans le systeme bi#2sr#2ca#1##yybycu#2o#8#+#x#+#y#/#2, montre que lorsque y est superieur a 0,5 les phases sont semiconductrices. L'utilisation de l'oxyde d'etain (sno) en tant qu'agent de cristallisation a permis d'obtenir des monocristaux de stchiometrie controlee. Les determinations structurales de deux des phases du systeme (ca, yb) montrent les modifications suivantes: decroissance de la longueur d'onde de la modulation, reorientation des paires electroniques non liees des atomes de bismuth, insertion d'atomes d'oxygene supplementaires entre les couches d'atomes de bismuth
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Bright, Colleen M. "'Knowing me - knowing you' : an exploratory and analytical study of the factors at an individual and organisation level which influence housing choices for older people." Thesis, University of South Wales, 2013. https://pure.southwales.ac.uk/en/studentthesis/knowing-me--knowing-you(46c5a99e-f8cc-4dcc-9e21-2150121adcbb).html.
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This research study is concerned with the projected, significant rise in the number of older people in the next 20 to 30 years with a consequent growth in demand for health, social care and specialised housing. There is also an added challenge of meeting this growing demand alongside government policy requirements on ‘choice’ and ‘voice’, as they operate in Wales, as well as current and projected fiscal challenges. In the context of the challenges posed by an increasing older people population this research study will explore the concept of choice and specifically whether housing choice exists for older people in maintaining their independence in old age. The significance of independent living in the context of this research study, relates to an individual’s ability to maintain choice and control over their daily lives within their home environment. The research findings indicate that decisions made, or not made, by individuals in terms of planning ahead for old age, can potentially impact on their ability to sustain independence as they age in the home of their choice. The tendency for older people to ‘discount the future’ is explored by the researcher highlighting the potential to influence such behaviour by encouraging older people to ‘count the future’ and plan for it in terms of their individual needs. The impact on individual housing choice of decisions made by organisations, in particular local authorities, the NHS and Registered Social Landlords, is also explored within the Literature Review and discussion of the research findings. The consequences of a growing elderly population will be explored by focusing on how the housing choices available to older people with a long term condition, and potentially increasing care needs as they age, may be influenced by decisions made at an individual and organisation level. The findings of this qualitative, exploratory study are based on data collected and analysed from 22 one to one, semi-structured interviews with 2 groups of older people. The Prospective Group (forward looking to potential changes in their housing needs) comprised 10 older people aged between 57 to 80 years, all of whom live in their own home in the community, and the Retrospective Group (looking back to the circumstances that prompted changes in their housing needs) comprised 12 older people aged between 66 to 84 years, all of whom have moved in to sheltered housing. The research findings were also reviewed and discussed with a number of community based groups and a Focus Group. Analysis and discussion of the research findings enabled the identification of a number of themes which the researcher has distilled in to 3 overarching themes: • Enabling informed choice • Issues for organisations • Issues for Individuals The findings of this research study are important because they illustrate that, while most if not all Individuals wish to remain independent in the home of their choice as they age, achievement of this outcome is usually left to chance. The researcher argues that this will continue to be the case unless action is taken at a national and local level to clarify the role of organisations and individuals in supporting and achieving independent living in to old age. The findings point to potentially significant implications for individuals and organisations in terms of an erosion of choices available for sustaining independence in to old age, at an individual level, and an inability to sustain services, at an organisation level. Emerging policy and continued national and local debate on the issues explored by this research study illustrate an increased focus on the consequences of an ageing population. The researcher suggests that future planning of housing and communities will need to more effectively reflect the diverse needs, wants and expectations of current and future generations of older people in terms of the homes they wish to live in.
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Wahbi, Mostafa. "Influence de l'ordre à courte distance sur les proprétés de conduction ionique de l'ion F- dans quelques solutions solides M2+1-XM'X2+alphaF21alphaX(alpha =1,2) de structure dérivée du type fluorine." Bordeaux 1, 1992. http://www.theses.fr/1992BOR10510.
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Differents processus d'ordre a courte distance ont ete mis en evidence et correles a la variation des proprietes electriques en fonction du taux de substitution au sein des solutions solides m#1##xbi#xf#2#+#x (m=sr, cd, ba, pb). La nature des ions fluorure interstitiels responsables des mouvements a longue distance dans ces series de materiaux a ete determinee. Les resultats ont ete obtenus a partir d'etudes par rmn du #1#9f et d'investigations par diffraction de neutrons et correles aux proprietes electriques par l'intermediaire du modele des processus de clusterisation. De nouveaux materiaux a base de pbf#2 et comportant deux cations substitutionnels (bi#3#+ et in#3#+, in#3#+ et zr#4#+, bi#3#+ et zr#4#+) ont ete prepares. Ces materiaux caracterises par la presence de clusters colonne de nature differente ont de tres bonnes conductivites ioniques, comparables a celle de pbsnf#4, meilleur conducteur anionique connu
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Coudour, Bruno. "Influence de la végétation et du relief dans les feux de forêt extrêmes : étude de l'accumulation, de la dégradation et des propriétés de combustion des composés organiques volatiles issus des feux de forêt." Thesis, Poitiers, 2015. http://www.theses.fr/2015POIT2290/document.
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Les pompiers méditerranéens sont confrontés à des embrasements soudains de la végétation (AFF) dont les mécanismes ne sont pas encore bien compris. La végétation étant l'unique combustible, nous nous sommes penchés sur les gaz qui en proviennent. Nous avons d’abord étudié la dégradation thermique de quatre Composés Organiques Volatils biogéniques (COVb) à l'aide d'une pyrolyse flash et d'un four tubulaire. À partir de cette étude et de la littérature, nous avons choisi un mélange d'étude afin expérimenter ses propriétés de combustion. Nous avons ainsi déterminé l'Énergie Minimale d’Inflammation (EMI) et la vitesse fondamentale de flamme de mélanges d'α-pinène/benzène qui sont respectivement les principaux COV détectés dans les plantes et dans les fumées de feux de forêt. Le dernier chapitre concerne l'étude stationnaire de l'accumulation de gaz dans des vallées à partir d'une maquette de forêt 1/400ème disposée dans une soufflerie<br>Mediterranean firefighters cope with powerful accelerations of forest fires (AFF) whose mechanisms are not very well understood. Vegetation is the only fuel of forest fire, then we studied the gases coming from them. First, we studied the thermal degradation of four Biogenic Volatil Organic Compounds (BVOCs) thanks to a flash pyrolysis and a tubular oven. From this study and literature, we chose a representative VOC mixture to study its combustion properties. We determined Minimal Ignition Energy (MIE) and its laminar burning speed of mixtures of α-pinene/benzene that are respectively the main VOC detected in vegetation and forest fire smoke. The last chapter experiment the steady-state gas accumulation above a 1/400 V-shaped forest model
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Vuong, Christine. "Evaluation of Sindbis-M2e Virus Vector as a Universal Influenza A Vaccine." Thesis, 2012. http://hdl.handle.net/1969.1/ETD-TAMU-2012-08-11706.
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Although avian influenza virus (AIV) infections in domestic poultry are uncommon, transmission of avian influenza from wild waterfowl reservoirs does occur. Depopulation of the infected flock is the typical response to AIV outbreaks in domestic chicken production, causing a loss in profits and accumulation of unexpected expenses. Because it is impossible to know which of many virus subtypes will cause an outbreak, it is not feasible for the U.S. to stockpile vaccines against all possible avian influenza threats. Currently, the U.S. does not routinely vaccinate chickens against influenza due to the inability to differentiate infected from vaccinated animals (DIVA), which would place limitations on its trade markets. A Sindbis virus vector expressing the PR8 influenza strain's M2e peptide was developed as a potential universal DIVA vaccine. M2e is a conserved peptide amongst influenza A viruses; M2e-specific antibodies induce antibody-dependent cytotoxicity or phagocytosis of infected cells, reducing production and shedding of AIV during infection. In this study, chickens were vaccinated at one-month-of-age with parental (E2S1) or recombinant Sindbis viruses expressing the PR8 M2e peptide (E2S1-M2e) by subcutaneous or intranasal routes at high (106 pfu) or low (103 pfu) dosages. Chickens were boosted at 2-weeks post-initial vaccination using the same virus, route, and dosage, then challenged with low pathogenic H5N3 AIV at 0.2 mL of 106/mL EID50 2-weeks post-boost. Serum samples were collected at 1-week and 2-weeks post-vaccination, 2-weeks post-boost, and 2-weeks post-challenge and screened for PR8 M2e-specific IgY antibody production by ELISA. Both high and low dose subcutaneously, as well as high dose intranasally vaccinated E2S1-M2e groups produced significantly higher levels of PR8 M2e-specific IgY antibodies as early as 1-week post-vaccination, while the uninoculated control and E2S1 groups remained negative for all pre-challenge time points. M2e-specific IgY antibodies capable of binding the challenge H5N3 M2e peptide were detected in groups with existing vaccine-induced M2e-specific antibodies pre-challenge, suggesting antibody M2e cross-reactivity. After challenge, all groups developed M2e-specific IgY antibodies and high HI titers, verifying successful AIV infection during challenge and production of hemagglutinin-specific antibodies. Viral shedding titers 4-days post-challenge were used to measure vaccine efficacy and were similar amongst all groups. Microneutralization assay results confirmed that post-boost serum samples, containing only M2e-specific antibodies, were unable to neutralize AIV in vitro. Although the E2S1-M2e vaccine was capable of producing high levels of M2e-specific IgY antibodies when inoculated subcutaneously, these antibodies were not able to reduce viral shedding and therefore did not protect chickens from AIV.
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Hasan, Noor Haliza. "Avian Influenza virus M2e protein: Epitope mapping, competitive ELISA and phage displayed scFv for DIVA in H5N1 serosurveillance." Thesis, 2017. http://hdl.handle.net/2440/119643.
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Within the avian influenza virus (AIV) history, H5N1 subtype is the most alarming in terms of its spread rate throughout the globe with its demonstrated unusual pattern of evolution. Persistency and constant circulation of this subtype in poultry population in a number of countries have resulted its establishment and declaration as enzootic. The affected countries are commonly characterised by high poultry populations and productions. They are also developing countries which have minimal funding allocated for precaution on disease incursion. Past observations showed that a single AIV epizootic is capable of causing significant economic burden throughout the world. Although epizootic, it still resulted sporadic cases of human infection and mortality. Therefore, H5N1 enzootic countries opt for vaccination strategy (usually with inactivated whole virus) to evade AIV incursions. However, this interferes with the AIV surveillance effort. This is due to the lack of diagnostic tool with the ability to differentiate AIV infected animal from vaccinated animal (DIVA). Following this realisation, several options are made available. Diagnostic tool development which is capable of DIVA requires a highly sensitive and specific target which at the same time is economic, and pose ease of application. In recent years, growing interest on the AIV matrix 2 extracellular domain (M2e) protein has propelled its exploration as the target for AIV serosurveillance diagnostic tool development. It has been demonstrated to be highly sensitive and specific in detection for AIV infection in an indirect enzyme-linked immunosorbent assay (ELISA) setting. The factor which made it highly interesting is its ability for DIVA application. M2e protein can only be found in low concentration on an AIV particle which is used in an inactivated vaccination strategy, while present in high concentration if cells are AIV infected. Therefore, this study has further explores the AIV M2e protein potential for AIV serosurveillance diagnostic tool development and successfully demonstrated an M2e-based test in a competitive ELISA format for DIVA. This particular ELISA format was of interest as it can be potentially used in multiple species application, as AIV is a multispecies pathogen. To ensure the universality of the competitor antibody, comparative mapping of anti-M2e antibodies from chicken, mouse and rabbit was done. Findings highlighted slight variations in the epitope identified for the M2e antigen by antibodies from different species. Mouse anti-M2e antibodies are more suitable to be used as the competitor antibodies against anti-M2e chicken sera in the M2e-based competitive ELISA test. Consequently, application of the mouse anti-M2e antibodies in the M2e-based competitive ELISA has demonstrated specific and sensitive indication of AIV infection in the H5N1 challenged chicken sera. Biotechnology developments has also introduced the single chain variable fragment (scFv) antibodies as specific and stable bait for antibodies detection against targeted pathogen’s protein (antigen). Taking advantage of this knowledge, this study has also successfully isolated reactive and specific anti-M2e scFv antibodies from avian sources. This is critical as an avian sourced antibodies to be used as bait for the targeted pathogen’s protein is highly relevant in the setting for AIV serosurveillance application in the poultry industry. These findings are significant in the effort to provide a highly sensitive and specific diagnostic tool, which are also cost effective, easy to apply with high throughput ability. Such ideal diagnostic tool for AIV serosurveillance is highly valuable, as this may hold the key to break the AIV continuous circulation.<br>Thesis (Ph.D.) -- University of Adelaide, School of Animal and Veterinary Sciences, 2017
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Chen, Jyun-Yu, and 陳浚宇. "RNF125 mediated influenza M2 protein degradation." Thesis, 2019. http://ndltd.ncl.edu.tw/handle/2deqzw.
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碩士<br>國立陽明大學<br>微生物及免疫學研究所<br>107<br>Influenza A virus belongs to Orthomyxoviridae, and is a common pathogen to human. Influenza viruses cause yearly outbreak around the world, resulting in about three to five millions cases of sever illness. Because the virus mutates frequently, we have to predict vaccine strains every year, moreover preexisting immunity cannot provide protection fully. Mutations also render influenza viruses resistance to current antivirals. A better understanding of host-virus interaction may helps us identify potential target for new antiviral drugs.
Activation of RIG-I-like Receptors ( RLR ) signaling pathway induces type I interferon production to defend against influenza infection. This signal is transmitted through the aggregation of the adaptor protein MAVS on mitochondria. Previously, we found that activation of RIG-I promoted protein degradation of influenza viral proteins. M2 and PB1-F2 were the most affected, so we try to understand the possible mechanism mediating their degradation. We have found that RNF125, an unbiquitin E3 ligase known to regulate RIG-I signaling negatively, played a role in RIG-I-induced PB1-F2 protein degradation. Here, we extended the study to M2. On the other hand, using different pharmacological inhibitors has previous helped us identified the autophagy may be part of RIG-I- induced viral protein degradation. In this study, we examined the role of autophagy in RNF125-mediated M2 degradation.
First, we found the overexpression of RNF125 facilitated the protein degradation of M2. Consistently, knockdown of RNF125 reduced RIG-I-induced M2 degradation. Next, by using pharmacological inhibitors, we found the M2 expression could be partially rescued by autophagy inhibitor including 3-MA and Bafilomycin-A1, suggesting M2 might be degraded through autophagy upon RIG-I activation. Surprisingly, we didn’t observe the proteasome inhibitors, such as MG132 and lactacystin, can reduce degradation. Finally, we investigated that whether RNF125 affects influenza virus ( PR8 ) during infection. We found that in presence of overexpressed RNF125, M2 expression was reduced moderately. This may lead to the reduction of viral tirter we observed. We concluded that RNF125 may play a role in RIG-I-mediatied antivral activity through enhancing viral protein degradation.
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Chen, Ying-Chun, and 陳盈均. "RIG-I mediated influenza M2 protein degradation." Thesis, 2017. http://ndltd.ncl.edu.tw/handle/45hw3r.
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碩士<br>國立陽明大學<br>微生物及免疫學研究所<br>105<br>RIG-I like receptors (RLR) signaling activation induces type I interferon response to defend against influenza virus infection. RIG-I and MAVS overexpression also induces autophagy to maintain mitochondrial homeostasis. In our previous studies, we observed that activation of RIG-I signaling led to PB1-F2 derived from A/Puerto Rico/34/8/H1N1 (PR8) degradation. As a result, we investigated whether activation of RIG-I signaling induces other influenza viral proteins degradation. We found that PR8 PB1, PA, HA and NP expression level decreased slightly when N-RIG, an active form of RIG-I, was overexpressed, while PR8 PB1-F2, M2 and NS2 viral proteins reduced more significantly. After comparing the expression level of different influenza viral proteins under N-RIG overexpression, we chose viral protein M2 as our model to study the degradation mechanisms. M2 is responsible for virus releasing to nucleus and is known to help virus to evade infection-induced autophagy by blocking autophagosome fusion with lysosome, thus M2 plays an important role in the life cycle of influenza virus.
To investigate how RIG-I mediated M2 degradation, cells were treated with different pharmacological inhibitors in the presence of RLR signaling activation. We observed that M2 expression level could be rescued by a proteasome inhibitor, lactacystin, and three autophagy-related inhibitors, 3-MA, Bafilomycin-A1 and Leupeptin, indicating that M2 might be degraded via both proteasomel and lysosomal pathways. In the study of autophagy and lysosomal pathways, we further observed that M2 could be degraded when cells were transfected with plasmids expressing EGFP-LC3 or treated with CCCP to induce autophagy. It showed that autophagy and lysosomal pathway indeed participated in RIG-I mediated M2 degradation. In the study of proteasome degradation pathway, we also investigated whether E3 ligases involved in RLR signaling led to M2 degradation. We found that several E3 ligases, such as RNF125, RNF135 and Parkin, could further enhance M2 degradation in the presence of RIG-I signaling, suggesting that E3 ligases might also play a role in RIG-I mediated M2 degradation. However, the detailed mechanisms remain to be deduced.
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廖若吟. "The immune response to Influenza A virus M2 protein." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/27124919320553852462.
Full textAbstract:
碩士<br>國立臺灣大學<br>醫事技術學研究所<br>90<br>Due to the antigenic variation of the viral surface proteins, hemagglutinin (HA) and neuraminidase (NA), the conventional influenza vaccine has to be evaluated every year. In order to improve the defectiveness, we chose another surface protein, M2 protein, to study and analyze the M2 protein specific immune response to understand if it is suitable as a vaccine component.
This thesis could be divided into two parts. The first part was to study the M2 antibody response after vaccination a commercial influenza vaccine, Pasteur vaccine. The inactivated vaccine was inoculated to human. Paired serum were collected and the M2 antibody was detected by Western blotting using recombinant M2 protein. The M2 specific antibody could not be detected in all of the serum. It was possible that the M2 protein present in the vaccine was too low to induce antibody response. High dose of the inactivated vaccine was inoculated into mice, no M2 specific antibody could be detected.
The second part is to study the immune response of M2 DNA vaccine. The mice were inoculated with three different M2 plasmids, and the M2 specific antibody was detected. It was found that the plasmid containing M2 (full) gene could induce M2 specific antibody six weeks after inoculation, and the highest antibody titer could be detected at the eighth week. To analyze the antibody subtype, we found that the major subtype was IgG2a. This implied a Th1 immune response. In the experiment to detect cellular immune response, the IFN-γ secreting cells were detected by ELISPOT. It was found that all of the three different M2 plasmids could induce cellular immune response. The neutralization assay showed that the M2 antibody could not neutralize the influenza virus.
Our results showed that no M2 specific antibody could be detected after inoculating inactivated influenza vaccine. However, the M2 specific antibody and cellular immune response could be detected after immunization with M2 (full) DNA vaccine. Further study of the protection efficiency in animals will be required to understand what the M2 protein could be one of the suitable vaccine components.
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"Japanese investment and influence in Thai development." The MIT Japan Program, Massachusetts Institute of Technology, 1991. http://hdl.handle.net/1721.1/17086.
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Li, Guan-Wei, and 李冠緯. "Self-assembly of bitopic membrane protein M2 of influenza A." Thesis, 2016. http://ndltd.ncl.edu.tw/handle/13847684437632686457.
Full textAbstract:
碩士<br>國立陽明大學<br>生醫光電研究所<br>104<br>Ion channels are pore-forming proteins in charge of permeability of specific ions on lipid bilayer by electrochemical gradients. They exist not only in every cellular membrane but in most of viruses and reveal the key part in how cells and viruses work.
To investigate how ion channel proteins interact with each other from the aspect of biophysics, there is a program built for helping the user to generate the whole energy profile. This program of ion channel assembly (PICA) has 5 degrees of freedom (DOFs), namely, D, θ r, θ t, N and h, in order to describe each of the ion channel structures. PICA screens every point of DOF in the range the user is setting. Unlike other existing docking program or software that they search space all around the target proteins for proper conformation, PICA only search the space around the side surface of trans-membrane domain (TMD) of protein to find the its possible pore structure. Thus, it is more like a 2 dimensional search method rather than 3 dimensional search algorithm like other programs for globular proteins.
The situation of PICA is different from those of the others tools. PICA is built under the assumption that pore-forming proteins are assembled on lipid bilayer of endoplasmic reticulum; that also is, these pore-forming proteins don’t need to be tilted at big angles and is assembled together under the lipid environment whose dielectric constant is nearly 2. An important feature of PICA is that the displacement of the structure includes the Cα atoms and sidechain. It turns out that the Root-Mean-Square Deviation (RMSD) values of top 10 structures are large; however, all of top 10 structures form proper orientation of residues to function as channels.
In this research, I also performed other docking software and programs to assemble M2 proton channel of influenza A virus. These software and programs are Molecular Operating Environment (MOE), Sam, M-ZDOCK and Rosetta. In order to see how well it is on predicting pore structures of membrane proteins, RMSD comparison of backbones between original structure and prediction is applied. I also checked the top 10 prediction structures which these software and programs provided to user.
Most of these prediction results failed to reproduce the actual structure of the M2 proton channel, and even though RMSD values of some of the predictions are small enough that they seem promising. In the case of M2 proton channel, the pore structure of the M2 channel must have four His and four Trp positioned inside the pore region in order to be proton conducting. The His of channel function as a pH sensor and the Trp forms the ion gate of the channel3. Most of the prediction results of existing software and programs have presented incorrect orientation of His and Trp.
The final comparison tells the users that PICA provided ten proper structures to the user, and Sam provided the two, and M-ZDOCK provided the one and both MOE and Rosetta failed to provide any proper structures on the docking of M2 proton channel of influenza A virus.