Dissertations / Theses on the topic 'Substrats interactifs'
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1
Gamouh, Ahmed. "Effets comparés et interactifs des pesticides et des facteurs physiques sur la minéralisation de divers substrats carbonés dans le sol." Perpignan, 1998. http://www.theses.fr/1998PERP0314.
Abstract:
Les objectifs de cette etude etaient (1) de permettre une meilleure interpretation des tests ecotoxicologiques bases sur la mesure de la mineralisation de substrats organiques marques au carbone 14 (glucose, acide benzoique, 2,4-d) par la microflore du sol, (2) de comparer les effets de pesticides (dnoc, aldicarbe) a ceux de facteurs physiques (congelation, sechage, chauffage), (3) d'estimer les effets combines des deux types de facteurs. L'etude a ete conduite selon un dispositif experimental simplifie. Dans un premier temps, les traitements physiques etaient appliques sur des echantillons de 2g de sol. Les substrats et les pesticides etaient ensuite introduits en solutions aqueuses. Les echantillons etaient incubes a 20c et la radioactivite degagee etait determinee apres 3h, 24h, 8j et 64j. Nos resultats ont confirme que le rapport entre les quantites de carbone mineralise et celles incorporees dans la biomasse microbienne augmente avec la concentration en substrat. En presence d'une forte biomasse et de concentrations faibles en substrat, les facteurs physique aussi bien que les pesticides peuvent provoquer une forte augmentation des proportions de carbone mineralise. En revanche, une faible biomasse et/ou de fortes concentrations en substrat ont pour consequence une diminution du niveau de co#2 degage. D'une facon generale, l'importance des effets augmente dans l'ordre : congelation, aldicarbe, sechage, chauffage a 50c et dnoc. Les effets des interactions entre facteurs physiques et chimiques sont generalement plus importants que chacun des effets individuels. D'une facon generale, l'importance des effets augmente dans l'ordre congelation, aldicarbe, sechage, chauffage a 50c et dnoc. Il existe de grandes similitudes entre les effets des traitements physiques et ceux des pesticides, ce qui pourrait permettre d'utiliser les premiers comme critere d'evaluation de l'impact ecologique des seconds. Le plus souvent l'interaction des deux types de facteurs produit un effet resultant plus important que celui de chaque effet individuel. Dans quelques cas on observe un effet synergique entre des facteurs physiques moderes (exemple la congelation) et les pesticides.
2
Battut, Alexandre. "Interaction substrates and instruments for interaction histories." Electronic Thesis or Diss., université Paris-Saclay, 2024. http://www.theses.fr/2024UPASG026.
Abstract:
Dans le monde numérique comme dans le monde physique, nos interactions avec les objets laissent des traces qui racontent l'histoire qui les a façonnés au fil du temps. Ces données historiques peuvent être consultées par les utilisateurs afin de mieux comprendre les étapes qui ont conduit à l'état actuel de leur système. Elles peuvent également être re-documentées afin d'arranger l'historique d'une manière plus compréhensible pour les utilisateurs. Dans des environnements collaboratifs, les utilisateurs peuvent être amenés à partager ces données, afin de mieux coordonner ou synchroniser leur travail d'équipe.Des travaux antérieurs ont tenté de démontrer les avantages des historiques partagés entre applications, mais les implémentations actu- elles des historiques dans les systèmes interactifs continuent de confiner les historiques à leur application d'origine.Les utilisateurs ne peuvent pas croiser leur historiques pour corréler les événements qui se sont produits dans différentes applications. Dans cette thèse, je montre que concevoir des historiques de l'interaction pouvant être partagés entre les applications et les utilisateurs faciliterait la navigation, la compréhension et la réutilisation des données historiques. J'ancre le début de mes travaux dans le cas de l'écriture collaborative afin d'explorer des écologies de traces et des usages familiers, mais néanmoins complexes. J'identifie les pratiques récurrentes et les problèmes liés à l'utilisation des données historiques en interrogeant des utilisateurs habitués de l'écriture collaborative, et je mène plusieurs activités de conception basées sur les observations qui en découlent. Je décris ensuite un premier système en tant que preuve de concept intégrant deux outils résultant de ces activités de conception. Ce système intègre également la première itération d'une structure unique pour les données d'historique partagées entre applications et utilisateurs. Les résultats des études utilisateurs menées sur ce système montrent que ces derniers expriment effectivement le besoin de disposer d'historiques d'interaction unifiés et personnalisables. En compilant les données recueillies au cours de ces activités de recherche et en me basant sur des travaux antérieurs concernant les "médias dynamiques partageables" et les substrats d'interaction, je décris un cadre permettant de concevoir des historiques d'interaction plus flexibles. Je présente Steps, une structure d'unification des données historiques qui intègre un noyau d'attributs descriptifs qui préserve l'intégrité d'une trace entre les applications, et des attributs contextuels extensibles qui permettent aux utilisateurs de modeler leurs historiques en fonction de leurs besoins. Je présente ensuite OneTrace, un prototype basé sur les Steps. Son implémentation suit mon cadre descriptif pour les historiques inter-applications et définit l'historique comme un matériau numérique à façonner par l'utilisation d'outils dédiés. Je discute des opportunités offertes par cette approche pour réaliser un changement de paradigme sur la façon dont nous concevons les historiques et leurs outilsIn the digital world, as in the physical world, our interactions with objects leave traces that tell the story of the actions that shaped these objects over time. This historical data can be accessed by end users to help them better understand the steps that led to the current state of their system. These traces can also be reused for activities such as re-documenting their own history to arrange it in a way that they find more understandable. Users may also be led to share these data in collaborative environments, to better coordinate and synchronize their work. While previous work has attempted to show the benefits of cross-application histories, current implementations of interaction histories in interactive systems tend to tie history data to their source application. This prevents users from cross-referencing historical data to review and correlate events that occurred in different applications.In this thesis, I argue that designing interaction histories that can be shared among applications and users would support browsing, understanding and reusing historical data. I first ground my work in the use case of collaborative writing to explore relatable yet complex traces ecologies and interaction history use. I identify recurring practices and issues with the use of history data by interviewing knowledge workers and conducting several design activities based on these observations. I describe a first proof-of-concept system integrating two history instruments resulting from these design activities, and the first iteration of a unifying structure for historical data to be shared among applications and users. The results of user studies show that users indeed express a need for unified and customizable interaction histories.Compiling the data gathered during these research activities and based on previous works about “Dynamic Shareable Media” and the Interaction Substrates and Instruments model, I describe a framework to help create more flexible interaction histories. The goal is to describe how to design interaction history systems that would help users take control of their historical data. I introduce Steps, a structure for unifying historical data that includes descriptive core attributes to preserve the integrity of a trace across applications, and extensible contextual attributes that let users reshape their histories to suit their needs. I then introduce OneTrace, a proof-of-concept prototype based on Steps that follows my descriptive framework for cross-application histories and defines interaction histories as digital material to be shaped by digital tool use. I discuss the opportunities offered by this approach to support a shift in paradigm on how we design and interact with interaction histories
3
Burns, Teresa Ellen. "Asymmetric Adsorbate and Substrate Interactions in Physisorbed Systems: N2 on Graphite and Dipolar Molecules on Ionic Substrates." DigitalCommons@USU, 1994. https://digitalcommons.usu.edu/etd/2090.
Abstract:
Asymmetries in physisorbed systems give rise to interesting phases and phase transitions in two-dimensional (2D) monolayer and multilayer systems. The effects of asymmetric adsorbate and substrate interactions in monolayers of dipolar molecules on ionic substrates and N2 on graphite are studied.
In the case of dipolar molecules on ionic substrates, 2D dielectric phase transitions using a modified Blume-Emery-Griffiths (BEG) model are determined theoretically. A dipole adsorbed vertically above a metal ion lattice site, and pointing up (down), is assigned a spin s=+1 (s=-1). An empty lattice site is assigned a spin S=0. Analytic solutions for both ferroelectrically and antiferroelectrically ordered systems are found. The model is applied to CO adsorbed on MgO and NaCl, and preliminary results for the phase diagram of CH3F on NaCl, are presented.
Multilayer phase transitions for N2 on graphite are studied experimentally using synchrotron x-ray diffraction. The system is measured to undergo layering transitions, where the number of layers increases as the temperature of the system increases. A new multilayer phase diagram based on our results and the combined results published by other researchers is presented. The effects of capillary condensation on this multilayer system are quantified, and it is determined that its primary effect is to broaden the discrete layering transitions. The results for both studies are put into context with other adsorption systems with asymmetric interactions.
4
Richter, Andreas. "Structure formation and fractionation in systems of colloidal rods." Phd thesis, Universität Potsdam, 2007. http://opus.kobv.de/ubp/volltexte/2007/1309/.
5
Sharp, Katherine Helen. "Substrate binding : interactions in ascorbate peroxidase." Thesis, University of Leicester, 2004. http://hdl.handle.net/2381/30091.
Abstract:
X-ray crystallography, kinetics, spectroscopy and directed evolution have been used to define substrate binding in recombinant soybean cystolic ascorbate peroxidase (rsAPX), which catalyses the hydrogen peroxide-dependent oxidation of ascorbate in plants.;The rsAPX crystal structure has been solved to 1.8 A (Chapter 2) and provides a useful comparison for the substrate bound structures. Nitric oxide and cyanide bound rsAPX crystal structures have also been solved to 2.0 A and are used to model the binding of hydrogen peroxide in APX (Chapter 2).;Ascorbate binding interactions are defined by the rsAPX/ascorbate crystal structure, which has been solved to 1.4 A (Chapter 3). This structure provides new rationalization of the unusual functional features of the related cytochrome c peroxidase enzyme, and a mechanism for electron transfer from the substrate to the heme has been prepared (Chapter 3).;The crystal structure of the rsAPX/salicylhydroxamic acid (SHA) complex has been solved to 1.46 A (Chapter 4). Spectroscopic and kinetic techniques were used to show that guaiacol and SHA bind at the same site in APX. These data define the aromatic binding site in rsAPX and, along with the other crystal structures, have been used to propose a mechanism for proton transfer (Chapter 4).;A screen for enhanced aromatic oxidation has been developed, using guaiacol (colourless), which is oxidised to tetraguaiacol (red) by rsAPX (Chapter 5). The colour change and the fact it can be carried out on whole cells, allows many mutants to be assessed very efficiently. As a result it will be ideal for screening libraries of rsAPX mutants created by directed evolution.
6
Jovelet, Cécile. "Interactions du vandetanib avec la P-glycoprotéine et passage d'une barrière physiologique : le placenta." Thesis, Paris 11, 2012. http://www.theses.fr/2012PA114825.
Abstract:
La surexpression de protéines d’efflux, et tout particulièrement la P-glycoprotéine, est impliquée dans la multidrug résistance. Dans cette thèse, nous démontrons que le vandetanib, inhibiteur de tyrosine kinase, est à la fois substrat et inhibiteur de la P-glycoprotéine et qu’il est capable de réverser in vitro la résistance à la doxorubicine liée à la surexpression de la P-glycoprotéine.Nous nous sommes également intéressés à l’étude du passage transplacentaire du vandetanib et nous montrons que ce médicament traverse la barrière placentaireOverexpression of ABC transporters, especially P-glycoprotein, is involved in multidrug resistance. In this study, we demonstrate that vandetanib, a tyrosine kinase inhibitor, is both substrate and inhibitor of P-glycoprotein and is able to reverse in vitro resistance to doxorubicin, linked to overexpression of P-glycoprotein.We also studied the placental transfer of vandetanib and we show that this drug crosses the placenta
7
Bischofs, Ilka Bettina. "Elastic interactions of cellular force patterns." Phd thesis, [S.l. : s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=973638915.
8
Hill, S. D. "Plasma torch interaction with a melting substrate." Diss., Georgia Institute of Technology, 1999. http://hdl.handle.net/1853/17261.
9
O'Donnell, Emily. "Substrate and redox partner interactions with CYP17." Thesis, University of Sheffield, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.427259.
10
Rahman, Nahid 1974. "Polypyrrole : an interactive substrate for bone regeneration." Thesis, Massachusetts Institute of Technology, 1998. http://hdl.handle.net/1721.1/50554.
Abstract:
Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Materials Science and Engineering, 1998.Includes bibliographical references (leaves 59-68).
Current methods of bone repair rely on autografts (bone from a donor site) and allografts (bone from human cadaver). However, these methods are plagued with disadvantages. There is a clear and urgent need to provide alternatives for regenerating and repairing bone. Bone is known to be one of the many connective tissues in the body that are responsive to exogenous electrical stimulation. Based on this principle, this thesis explores the potential of using an electrically conducting polymer, polypyrrole, as a substrate for bone regeneration. Optically transparent thin films of polypyrrole, with a polyanionic dopant, poly(styrenesulfonate), were synthesized electrochemically and characterized by X-Ray Photoelectron Spectroscopy, UV/VIS spectroscopy, Scanning Electron Microscopy and by electrical conductivity measurements. In this study, Bone Marrow Stromal Cells (BMSC), which are the progenitor cells to bone cells (osteoblasts), were used as the in vitro model system. Their viability, proliferation and differentiation capabilities were evaluated on polypyrrole, in the absence and presence of electrical stimulation. Results indicate that polypyrrole is ideally suited as a substratum for BMSC growth and differentiation. The application of an electrical stimulus through the polypyrrole substrate was found to induce the differentiation of BMSC towards an osteogenic lineage. Thus, polypyrrole, by virtue of its conductive properties, its in vitro biocompatibility and its flexibility in altering surface characteristics, has an exciting potential as a suitable interactive substrate for bone regeneration.
by Nahid Rahman.
S.M.
11
Wöbking, Barbara. "Substrate interactions on the ABC transporter MsbA." Thesis, University of Cambridge, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.613410.
12
Rahman, Nahid S. M. Massachusetts Institute of Technology. "Polypyrrole : an interactive substrate for bone regeneration." Thesis, Massachusetts Institute of Technology, 1998. http://hdl.handle.net/1721.1/50554.
Abstract:
Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Materials Science and Engineering, 1998.Includes bibliographical references (leaves 59-68).
Current methods of bone repair rely on autografts (bone from a donor site) and allografts (bone from human cadaver). However, these methods are plagued with disadvantages. There is a clear and urgent need to provide alternatives for regenerating and repairing bone. Bone is known to be one of the many connective tissues in the body that are responsive to exogenous electrical stimulation. Based on this principle, this thesis explores the potential of using an electrically conducting polymer, polypyrrole, as a substrate for bone regeneration. Optically transparent thin films of polypyrrole, with a polyanionic dopant, poly(styrenesulfonate), were synthesized electrochemically and characterized by X-Ray Photoelectron Spectroscopy, UV/VIS spectroscopy, Scanning Electron Microscopy and by electrical conductivity measurements. In this study, Bone Marrow Stromal Cells (BMSC), which are the progenitor cells to bone cells (osteoblasts), were used as the in vitro model system. Their viability, proliferation and differentiation capabilities were evaluated on polypyrrole, in the absence and presence of electrical stimulation. Results indicate that polypyrrole is ideally suited as a substratum for BMSC growth and differentiation. The application of an electrical stimulus through the polypyrrole substrate was found to induce the differentiation of BMSC towards an osteogenic lineage. Thus, polypyrrole, by virtue of its conductive properties, its in vitro biocompatibility and its flexibility in altering surface characteristics, has an exciting potential as a suitable interactive substrate for bone regeneration.
by Nahid Rahman.
S.M.
13
Zhang, Baoshe. "A study of substrate--liquid crystal interaction /." View Abstract or Full-Text, 2003. http://library.ust.hk/cgi/db/thesis.pl?PHYS%202003%20ZHANG.
Abstract:
Thesis (Ph. D.)--Hong Kong University of Science and Technology, 2003.Includes bibliographical references (leaves 176-186). Also available in electronic version. Access restricted to campus users.
14
Layton, Stephanie Danielle. "Identifying Kinase/Substrate Interactions in Chlamydia trachomatis." OpenSIUC, 2014. https://opensiuc.lib.siu.edu/theses/1575.
Abstract:
Chlamydia spp. are obligate intracellular, gram negative bacterial pathogens responsible for a variety of diseases in humans and animals. While vaccines are available for C. felis and C. abortus, they are not available for other chlamydial species. Antibiotics are commonly used to treat chlamydial infections, but resistance has been seen in some species, and low dosage can lead to persistence. These problems support the need for further research into factors mediating chlamydial infections, which may identify novel therapeutic targets and/or vaccine strategies. Protein phosphorylation on serine, threonine, and/or tyrosine residues has been identified in prokaryotes (performed by Hanks-type kinases), and these modifications serve important roles in development and virulence. Three genes with homology to Hanks-type kinases were found when analyzing the C. trachomatis serovar D genome (PknD, Pkn1, and Pkn5). Amino acid alignments were performed to compare the similarity of these three kinases between nine different chlamydial species. To identify kinase-substrate pairings, the Bacterial Adenylate Cyclase Two-Hybrid (BACTH) system was used to test putative kinase substrates along with development of a C. trachomatis BACTH expression library. To further validate kinase substrate pairings and to perform kinase inhibitor screens, a kinase assay was developed. Mutant kinases were generated to validate kinase activity. Collectively, this work has built a framework for directed and library-based kinase-substrate interaction testing and for high throughput kinase activity analysis to aid in inhibitor discovery.
15
Cubaka, Alfred. "Interactions plantes-bactéries sur des substrats contaminés en cuivre." Doctoral thesis, Universite Libre de Bruxelles, 2010. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/210068.
Abstract:
En utilisant le binôme Cupriavidus metallidurans CH34-Solanacées comme un modèle et comme un point dedépart, une étude sur l'interaction entre les plantes et les bactéries sur un substrat pollué par le cuivre a été menée
dans deux directions:
1 °) une étude en conditions de laboratoire sur les capacités de C. metallidurans CH34 à interagir avec Nicotiana
plumbaginifolia (les solanacées)
2 °) une étude sur le terrain visant à examiner les interactions entre cuprophytes et bactéries résistantes aux
métaux des régions minières du Katanga.
La première partie inclut une étude in silico visant à établir un catalogue des gènes C. metallidurans CH34
potentiellement impliqués par les interactions plantes-bactéries. Ce catalogue, tout en se reposant sur le génome
proche de Ralstonia solanacearum, bactérie phytopathogène de plusieurs espèces végétales appartenant
principalement à la famille des Solanaceae, il n'a pas pris en compte les orthologues des gènes clés de la
virulence de cette phytopathogène. Les gènes correspondants de C. metallidurans étaient situées sur les deux
chromosomes et ont des orthologues dans tous les génomes séquencés des Cupriavidus / Ralstonia et dans
Enterobacter sp. 638, endophyte de peuplier. L'étude transcriptomique, à l'aide de «microarray» a montré que
certains de ces gènes étaient induits, notamment des gènes impliqués dans la mobilité flagellaire (comme motA)
et dans la synthèse des polysaccharides extracellulaires étaient surexprimés pendant le contact entre les plantes et
les bactéries, tandis que phcA (impliqué dans la détection de la densité de population et dans la conversion
phénotypique) et des gènes impliqués dans la biosynthèse de pili étaient sousexprimés dans les conditions
expérimentales testées. En outre, le contact avec les plantes semble avoir induit la surexpression des gènes
impliqués dans la réponse de cuivre et d'autres métaux. La capacité de C. metallidurans CH34 à coloniser
l'endosphere de N. plumbaginifolia a été confirmée in vitro ainsi qu'un effet de promotion de la croissance des
plantes dans certaines conditions. Mais la densité de la colonisation (104-106 c.f.u/g. poids frais) est
considérablement réduite dans des conditions non stériles et en l'absence de pression de sélection métallique.
La deuxième partie de l'étude s'est concentrée sur la microbiologie de cuprophytes (Haumaniastrum katagense et
Crepidorhopalon tenuis) dans l'arc cuprifère du Katanga: des isolats Cuprorésistants appartenant aux genres
Stenotrophomonas et Sphingomonas prédominent dans la rhizosphère alors que des isolats appartenant aux
genres Methylobacterium, Xanthomonas et Variovorax prédominent dans l'endosphere. Certaines de ces
bactéries sont plus résistantes au Cu(II), à des concentrations minimales inhibitrices (MIC) allant jusqu'à 5 mM,
que C. metallidurans CH34 (MIC: 1,5 mM) et la plupart d'entre elles résistent également aux Zn(II), Co(II) et
Cd(II). Des isolats appartenant au genre Cupriavidus/Ralstonia ont été détectés dans la rhizosphère des
cuprophytes ainsi que les séquences 16S rDNA de C. metallidurans ont été également détectées dans l'ADN
total extrait des cuprophytes. La détection via la réaction de la polymérase en chaîne (PCR) de gènes de
résistance au cuivre correspondant à des protéines periplasmiques a confirmé la présence dans les bactéries
cuprorésistantes, principalement de copA et dans une moindre mesure celle de copK. Mais les gènes homologues
de copA et de copK n'ont pas été détectés dans tous les bactéries du genre Methylobacterium dont les membres
ont été pourtant les plus résistants aux métaux. Certaines bactéries isolées sont capables d'interagir avec le
système hormonal végétal et quelques unes semblent également manifester un effet de promotion de la
croissance des plantes. Les premières tentatives d'élaboration de protocoles de reinoculation des bactéries
endophytic cuprorésistantes dans Haumaniastrum katagense ont été effectués. La biologie moléculaire et
l'écologie des interactions plantes-bactéries et des mécanismes de résistance métallique décrits dans ce travail
peuvent préparer la voie à de nouvelles applications en bioremédiation (phytostabilization / phytoextraction de
métaux toxiques).
Doctorat en Sciences
info:eu-repo/semantics/nonPublished
16
Linenberger, Kimberly J. "Biochemistry Students' Understandings of Enzyme-Substrate Interactions as Investigated through Multiple Representations and the Enzyme-Substrate Interactions Concept Inventory." Miami University / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=miami1321309534.
17
Qi, Xiaolin. "Enzyme-substrate interactions in PC1 #beta#-lactamase catalysis." Thesis, University of Newcastle Upon Tyne, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315617.
18
Marsden, Catherine Jane. "Interactions between A-chain and it ribosomal substrate." Thesis, University of Warwick, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.269188.
19
Willenbockel, Martin [Verfasser], Frank Stefan [Akademischer Betreuer] Tautz, and Petra [Akademischer Betreuer] Tegeder. "Interacting Interactions: A Study on the Interplay of Molecule-Molecule and Molecule-Substrate Interactions at Metal-Organic Interfaces / Martin Willenbockel ; Frank Stefan Tautz, Petra Tegeder." Aachen : Universitätsbibliothek der RWTH Aachen, 2014. http://d-nb.info/1127861239/34.
20
Arjmandi-Tash, Omid. "Interaction of droplets and foams with solid/porous substrates." Thesis, Loughborough University, 2017. https://dspace.lboro.ac.uk/2134/24889.
Abstract:
Current problems on the interaction of complex liquids (i.e. droplets or foams) with complex surfaces (i.e. soft deformable or porous surfaces) are addressed in the following areas: (1) wetting of deformable substrates and surface forces, (2) kinetics of wetting and spreading of non-Newtonian liquids over porous substrates, (3) kinetics of spreading of non-Newtonian solutions over hair, (4) free drainage of foams produced from non-Newtonian solutions, and (5) foam drainage placed on porous substrates. Equilibrium of liquid droplets on deformable substrates was investigated and the effect of disjoining pressure action in the vicinity of the apparent three phase contact line was taken into account. It was proven that the deformation of soft solids is determined by the action of surface forces inside the transition zone. Spreading/imbibition of blood, which is a power law shear thinning non-Newtonian liquid, over a dry porous layer was investigated from both theoretical and experimental points of view. It was found that blood droplet spreading/imbibition over porous substrates shows two different behaviours: (i) partial wetting case with three subsequent stages: initial fast spreading, constant maximum droplet base and the shrinkage of the droplet base; (ii) complete wetting case with only two stages: initial fast spreading and the shrinkage of the droplet base. The wetting of hair tresses by aqueous solutions of two commercially available polymers, AculynTM 22 (A22) and AculynTM 33 (A33) was investigated experimentally. Both A22 and A33 solutions demonstrate well pronounced shear thinning behaviour. Initial contact angle of the A22 and A33 solutions on hair tresses was about 100o. The A22 droplets remained on the hair tress after spreading for at least half an hour. However, a fast penetration of the A33 droplets inside the hair tresses was observed when advancing contact angle in the course of spreading reached a critical value of about 60o. This could be explained by Cassie-Wenzel wetting transition which is caused by filling the pores inside the porous media by liquid. The influence of non-Newtonian rheology of A22 and A33 solutions on foam drainage was also investigated experimentally and a new theory of foam drainage was presented for the case of free drainage. For lowly viscous polymeric solutions and under the assumption of rigid surface of the Plateau border, the predicted values of the time evolution of the foam height and liquid content were in good agreement with the experimental data. However, in the case of highly viscous solutions an interfacial mobility at the surface of the Plateau border has to be taken into account. A completely new theory of foam drainage placed on porous substrate was developed. It was found that there are three different regimes of the process: (i) a rapid imbibition, the imbibition into the porous substrate dominates as compared with the foam drainage; (ii) an intermediate imbibition, that is, the imbibition into the porous substrate and the rate of drainage are comparable; (iii) a slow imbibition, the rate of drainage inside the foam is higher than the imbibition into the porous substrate for a period of time and a free liquid layer is formed over the porous substrate.
21
Douzi, Badreddine. "La machinerie de sécrétion de type II Xcp de Pseudomonas aeruginosa : relations structure-fonction et interactome." Thesis, Aix-Marseille 1, 2011. http://www.theses.fr/2011AIX10086.
Abstract:
Les bactéries à Gram négatif sont entourées par une enveloppe cellulaire qui, contrairement aux bactéries à Gram positif, possèdent une organisation membranaire complexe composée d’une membrane interne appelée généralement membrane cytoplasmique, un espace périplasmique contenant une matrice de peptidoglycane et une membrane externe asymétrique constituée d’une monocouche de phospholipides surmontée d’une assise de lipopolysaccharide (LPS). Afin de franchir cette barrière, les bactéries à Gram négatif ont développé différentes voies de sécrétions spécifiques dédiées à l’export des protéines (effecteurs) du milieu intracellulaire vers le milieu extracellulaire. Jusqu'à présent, six systèmes de sécrétion ont été identifiés chez ces bactéries. Chez Pseudomonas aeruginosa, une bactérie pathogène opportuniste, le système de sécrétion de type II appelé aussi sécréton Xcp constitue l’un des facteurs principales de sa virulence. Le sécréton Xcp est un complexe macromoléculaire formé par 12 protéines, nommées XcpAO et XcpPC-XcpZM. Ce complexe macromoléculaire est organisé en trois sous-complexes : i) une plateforme d’assemblage ancrée dans la membrane interne formé par les protéines XcpRESFYLZM ii) un pore de sécrétion localisé dans la membrane externe formé par l’oligomérisation d’une protéine appelé la sécrétine XcpQD. Le pore de sécrétion est connecté à la plateforme de la membrane interne par une protéine appelée XcpPC iii) un pseudopilus périplasmique sous forme de fibre hélicoïdale qui est formé par la multimérisation d’une protéine appelée la pseudopiline majeure XcpTG. D’autres protéines appelées les pseudopilines mineures XcpUH-VI-WJ-XK intègrent le pseudopilus. La première partie du travail effectué au cours de cette thèse a eu pour but d’étudier et de comprendre par des approches structurales, biochimiques et biophysiques le mécanisme d’assemblage des pseudopilines en pseudopilus. La deuxième partie de ce travail a porté sur l’étude des réseaux d’interactions entre les substrats sécrétés et les composants de la machinerie Xcp. Durant cette thèse, nous avons ainsi i) identifier grâce à l’étude des interactions protéine-protéine l’existence d’un complexe quaternaire entre les pseudopilines mineures XcpUH-VI-WJ-XK localisées au sommet du pseudopilus ii) déterminer les structures de la pseudopiline majeure XcpTG par RMN et de la pseudopiline mineure XcpWJ par cristallographie aux rayons X iii) déterminer les différents éléments du sécréton qui interagissent avec les exoprotéines du sécréton. Ce réseau d’interaction nous a permis de proposer un modèle de fonctionnement du sécréton qui élucide le cheminement des exoprotéines dans le sécréton afin qu’elles soient exportées vers le milieu extracellulaireGram-negative bacteria are characterized by a complex organization of their cell envelope composed by the inner membrane (IM) called cytoplasmic membrane, the periplasmic space containing a peptidoglycan layer and the outer membrane (OM) covered by the lipopolysaccharide matrix. Gram-negative bacteria have evolved several specialized machines called secretion systems to export their effectors from the intracellular medium to the extracellular milieu or to the host cells. Up to now, at least six secretion systems have been identified. In the opportunistic pathogen Pseudomonas aeruginosa, the type II secretion system called the Xcp secreton is the major pathway for the release of virulence factors. The Xcp secreton is a macromolecular complex composed by 12 proteins called XcpAO, XcpPC-XcpZM. This machinery is organized in 3 sub-complexes: i) the assembly platform localized in the IM implicating XcpRESFYLZM proteins ii) the OM pore composed by the oligomerization of the secretin XcpQD. The connection between the assembly platform and the secretin is performed by XcpPC anchored in the IM iii) a periplasmic pseudopilus consisting of the multimerization of the so-called major pseudopilin XcpTG. The pseudopilus is a helicoidally filament spanning the periplasmic area and pushing the substrate into the secretin pore. Four other proteins, the minor pseudopilins XcpUH-VI-WJ-XK, were found in the pseudopilus. In the present work we first focused on the study of the pseudopilus components by biochemical, biophysical and structural strategies to understand their assembly. Secondly, we investigate the protein interactome between periplasmic secreton component and secreted substrates. Thus, we revealed the presence of a quaternary complex composed by XcpUH-VI-WJ-XK located at the tip of the pseudopilus. To understand at atomic scale the regulation of the pseudopilus, we determined the structure of two components of the pseudopilus XcpTG by NMR and XcpWJ by X-ray crystallography. Using systematic protein-protein interaction studies between secreton components and purified exoproteins of Pseudomonas aeruginosa, we identified 5 proteins of the secreton able to interact with exoproteins. This interaction network allowed us to propose a model for the secretion process including the sequential steps followed by exoproteins inside the secreton to leave the cell envelop
22
Pauthe, Emmanuel. "Approches cinétiques et moléculaires de la reconnaissance enzyme-substrat : application à l'étude de l'activité protéolytique de la thermolysine." Compiègne, 1998. http://www.theses.fr/1998COMP1139.
Abstract:
L’accomplissement de tout acte protéolytique implique nécessairement la formation d'un complexe entre l'enzyme et son substrat. Par différentes approches cinétiques, spectroscopiques et moléculaires nous avons cherché à caractériser les phénomènes mis en jeu au cours de l'hydrolyse, par la thermolysine, de petits peptides en milieu biomimétiques. Cette étude a été conduite à l'interface entre la biochimie, la biophysique, la chimie et la physique. Dans un premier temps, nous nous sommes intéressés au comportement catalytique de la thermolysine sur des substrats modèles et en milieu modifié. Nous avons montré d'une part, que l'ajout d'additifs polyhydroxylés influence grandement l'activité de la thermolysine et d'autre part, affine les connaissances sur la spécificité et la sélectivité de cette enzyme (en particulier, mise en évidence de l'influence du résidu P'2 dans le mécanisme). Dans un deuxième temps, nous présentons des études structurales des peptides substrats en milieu modifié. Nous avons mis en évidence l'absence d'influence du micro-environnement contenant une forte proportion de glycérol sur la conformation des molécules de substrat et le rôle possible de leur structure tridimensionnelle quant à leur hydrolyse. Ces études ont été étendues à un autre modèle peptidique, de forme cyclique ou linéaire, et corrélées aux résultats cinétiques. Dans un troisième temps, par deux approches différentes, nous avons abordé l'étude des relations structure-fonction de la thermolysine. Expérimentalement, par des études cinétiques avec l'enzyme immobilisée et des déterminations de sa structure par spectroscopie laser Raman, nous montrons que l'enzyme est très peu sensible au micro-environnement. Théoriquement en analysant, par modélisation, l'interaction de la thermolysine avec un tripeptide substrat, nous avons mis en évidence des changements de conformation du substrat et/ou des mouvements du site actif enzymatique au cours de l'acte catalytique.
23
Schmidt, Matthieu. "Substrats neurophysiologiques des interactions patient- ventilateur et des sensations respiratoires correspondantes." Thesis, Paris 6, 2014. http://www.theses.fr/2014PA066487/document.
Abstract:
En ventilation assistée, l’inadéquation entre l’activité des muscles respiratoires du patient et l’assistance délivrée par le ventilateur se traduit par la survenue d’une dysharmonie patient-ventilateur potentiellement associée avec la survenue d’asynchronies patient-ventilateur et d’une dyspnée. Minimiser cette dysharmonie est un objectif majeur de la ventilation assistée. Le Neuro Asservissement de la Ventilation Assistée (NAVA) et la Ventilation Assistée Proportionnelle (PAV) sont deux nouveaux modes qui pourraient améliorer l’harmonie patient-ventilateur. Nous avons montré que, de façon similaire, le NAVA et la PAV diminuent le nombre d’asynchronie patient-ventilateur, préviennent la surdistension pulmonaire, restaurent la variabilité cycle à cycle du comportement ventilatoire et améliorent l’équilibre charge-capacité et le couplage neuromécanique. De plus, l’utilisation du mode NAVA en ventilation non invasive pourrait également permettre d’améliorer la synchronisation patient-ventilateur. Nous avons également montré aux cours de différents travaux sur la dyspnée en ventilation mécanique que celle ci était fréquente mais néanmoins difficile à identifier, en particulier chez les patients non communicants. L’EMG de surface des muscles inspiratoires extra-diaphragmatiques pourrait constituer un outil simple et objectif pouvant permettre au clinicien de diagnostiquer une dyspnée en ventilation mécanique et optimiser les réglages du ventilateur dans le but de minimiser la dysharmonie patient-ventilateur. Ces données permettent de progresser vers une meilleure connaissance de la dysharmonie patient- ventilateur. L’impact clinique de l’utilisation des modes proportionnels et d’une détection précoce de la dyspnée doit maintenant être évalué par des essais cliniquesVentilatory support must be tailored to the load capacity balance of the respiratory system to avoid patient-ventilator dysharmony as it may lead to patient-ventilator asynchronies and dyspnea. Minimizing this dysharmony is crucial. Neurally Ventilatory Assist Ventilation (NAVA) and Proportional Assist Ventilation (PAV) modes may improve patient-ventilator interaction. We showed in this work that PAV and NAVA both prevents overdistension, restores breath by breath variability of the breathing pattern and improves neuromechanical coupling and patient- ventilator asynchrony in fairly similar ways compared to pressure support ventilation. In addition the use of NAVA with non-invasive ventilation may also improve patient-ventilator interaction. We also demonstrated that dyspnea is a frequent issue in mechanically ventilated ICU patients and it can be difficult to assess when the patient is unable to report it. Surface electromyograms of extradiaphragmatic inspiratory muscles provides a simple, reliable and non-invasive indicator of respiratory muscle loading/unloading in mechanically ventilated patients. Because this EMG activity is strongly correlated to the intensity of dyspnea, it could be used as a surrogate of respiratory sensations in mechanically ventilated patients, and might, therefore, provide a monitoring tool in patients in whom detection and quantification of dyspnea is complex if not impossible. These data provide a better understanding of patient-ventilator dysharmony. Further studies are needed to evaluate the possible clinical benefits of NAVA and PAV on clinical outcomes and the impact of an early detection of dyspnea in mechanical ventilation
24
Ulrich, Magdalena Maria Wilhelmina. "Enzyme/substrate interactions of the vitamin K-dependent carboxylase." [Maastricht : Maastricht : Rijksuniversiteit Limburg] ; University Library, Maastricht University [Host], 1991. http://arno.unimaas.nl/show.cgi?fid=6207.
25
Hodge, Thomas C. "Substrate-film interaction in noble metal/polymer multichip modules." Thesis, Georgia Institute of Technology, 1996. http://hdl.handle.net/1853/10972.
26
Eggenhuisen, Joris Theodoor. "'The interaction between substrate evolution and turbidity current development'." Thesis, University of Leeds, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.507691.
27
Kinstrie, Ross Stuart. "Identification of Drosophila DYRK family substrates and interacting proteins." Thesis, University of Glasgow, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.433084.
28
Riley, Jane. "The interaction of topoisomerase IV with potential DNA substrates." Thesis, University of Liverpool, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.272768.
29
Hersch, Greg Louis. "ClpX interactions with ClpP, SspB, protein substrate and nucleotide." Thesis, Massachusetts Institute of Technology, 2005. http://hdl.handle.net/1721.1/34199.
Abstract:
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, February 2006.Includes bibliographical references.
ClpXP and related ATP-dependent proteases are implements of cytosolic protein destruction. They couple chemical energy, derived from ATP hydrolysis, to the selection, unfolding, and degradation of protein substrates with the appropriate degradation signals. The ClpX component of ClpXP is a hexameric enzyme that recognizes protein substrates and unfolds them in an ATP-dependent reaction. Following unfolding, ClpX translocates the unfolded substrate into the ClpP peptidase for degradation. The best characterized degradation signal is the ssrA-degradation tag, which contains a binding site for ClpX and an adjacent binding site for the SspB adaptor protein. I show that the close proximity of these binding elements causes SspB binding to mask signals needed for ssrA-tag recognition by ClpX. The SspB dimer overcomes this signal masking by tethering itself and bound substrate to ClpX, via docking sites located in the dimeric N-terminal domain of ClpX. Because this N-domain dimer binds only a single SspB subunit, the ClpX hexamer can accommodate just one SspB dimer per hexamer. Other adaptor proteins that use these same tethering sites must compete with SspB for access to ClpXP. Substrates bearing ssrA tags with increased spacing between the SspB and ClpX binding elements are degraded more efficiently at low concentrations by ClpXP.
(cont.) This mechanism in which the adaptor first obstructs and then stimulates substrate recognition may have evolved to permit an additional level of regulation of substrate choice. SspB binding to ssrA-tagged substrate is a highly dynamic process, allowing rapid transfer of substrates from SspB to ClpX. Although the ClpX hexamer is composed of six identical polypeptides, individual subunits assume at least three distinct conformations. Using a hexamer that was engineered to prevent nucleotide hydrolysis, I show that some nucleotide-binding sites in ClpX release ATP rapidly, others release ATP slowly, and at least two sites remain nucleotide free. Occupancy of both the slow sites by ATP and the fast sites by either ATP or ADP is required to bind the degradation tags of protein substrates. The ability of ClpX to retain binding of substrate with ATP or ADP in the fast sites suggests that nucleotide hydrolysis in the fast sites, but not in the slow sites, will allow repeated unfolding attempts without substrate release over multiple ATPase cycles. My results rule out ATPase models including ClpX6eATP6 or ADP6 and also suggest that the enzyme hydrolyzes only a fraction of bound ATP in a single turnover event. Short peptide motifs of ClpX, known as IGF loops, interact with ClpP and change conformation as a response to nucleotide binding by ClpX.
(cont.) As ClpX varies its nucleotide content during the ATP hydrolysis cycle, it also varies its affinity for ClpP. Processing of substrates is coupled to the ATP-hydrolysis cycle of ClpX and appears to modulate ClpX's affinity for ClpP by changing how long each ClpX subunit spends in each nucleotide state.
by Greg Louis Hersch.
Ph.D.
30
Jones, Darran Dafydd. "Protein-domain interactions and substrate channelling in multienzyme complexes." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621518.
31
Gullekson, Corinne. "Effect of Cell-Substrate Interactions on Epithelial Cell Mechanics." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/38174.
Abstract:
Cell-substrate interactions play a key role in the regulation of epithelial cell mechanics. Through a series of studies, we demonstrate how substrate interactions impact both the response to an oncogene and the cellular contractility and organization of a monolayer. We first examine the effects of oncogenic Ras in cells in adherent and suspended states.
To accomplish this, we utilized atomic force microscopy and a microfluidic optical stretcher. We found that adherent cells stiffen and suspended cells soften with the expression of constitutively active Ras. The effect on adherent cells was reversed when contractility was inhibited with the ROCK inhibitor Y-27632, resulting in softer cells. These findings suggest that increased ROCK activity as a result of Ras has opposite effects on suspended and adhered cells. In a subsequent study, we examined the effects of a substrate on contracting and relaxing monolayers. We created a new methodology for measuring the mechanodynamics of epithelial monolayers by culturing cells at an air-liquid interface.
These model monolayers were grown in the absence of any supporting structures in hanging drops. We found that the direction of strain in the unsupported monolayers was not correlated to nuclear alignment as observed when the monolayers were grown on soft deformable gels. It was also observed that both gel and glass substrates led to the promotion of long-range cell nuclei alignment not seen in the unsupported monolayers. To further characterize the morphology and mechanics of monolayers clusters observed in our experiments, we created a new computational model based on the vertex model. The energy function used in this model takes into account cell-cell and cell-substrate adhesion as well as anisotropic cellular mechanical properties. The results of these simulations suggest that the promotion of long-range alignment on solid substrates were due to cells having anisotropic elastic moduli with global alignment. They also suggest that the alignment observed in monolayers grown on air water interfaces is due to cells having low substrate adhesion and isotropic moduli. Our findings establish the importance of studying epithelial cell mechanics in different states of attachment.
32
Leckey, Jill J. "Exercise-nutrient interactions: Effects on substrate metabolism and performance." Thesis, Australian Catholic University, 2017. https://acuresearchbank.acu.edu.au/download/8abaa0b1b76c3014cfc2aad93fc50c5008eef9e6b6e8bbe177c3a02c38715a1e/5618185/Leckey_2017_Exercise_nutrient_interactions_effects_on_substrate_metabolism.pdf.
Abstract:
During prolonged (> 90 min), continuous steady-state exercise, skeletal muscle is fuelled by both carbohydrate (CHO) (i.e. muscle and liver glycogen, blood glucose and muscle, blood and liver lactate) and fat substrates (i.e. adipose and intramuscular triglycerides [IMTGs], blood-borne free fatty acids [FFAs] and TGs). The specific pattern of substrate oxidation is influenced by the relative exercise intensity, an individual’s training status and their preceding diet. However, it is well accepted that when exercising at high relative intensities (i.e. > 70% maximal oxygen uptake [V̇ O2max]), CHO-based fuels are the predominant fuel source. Despite CHO being important for sustaining prolonged exercise, recent attention has focused on exercise-nutrient protocols that reduce skeletal muscle dependence on CHO fuel sources and, instead, increase reliance on fat-based fuels. Such strategies include high-fat, low-CHO diets, training with low endogenous and exogenous CHO availability and oral ketone supplementation. In theory, strategies that “spare” the oxidation of CHO substrates should enhance endurance exercise performance. This thesis comprises a series of independent but related studies investigating the effects of manipulating both endogenous and exogenous fat availability on substrate metabolism, skeletal muscle adaptations and exercise performance.
Study 1 (described in chapter 4) investigated the effect of decreasing circulating FFA availability prior to and during half-marathon running. FFA availability was suppressed via the administration of nicotinic acid, ingested prior to and during exercise. The suppression of lipolysis and the exercise-induced rise in plasma FFAs did not impair half-marathon running capacity. When running at ~80% V̇ O2max for ~90 min there was a small but obligatory use of fat substrates, independent of CHO intake pre- and during exercise. However, CHO was the predominant fuel source, contributing between 80-90% to total energy expenditure.
Study 2 (described in chapter 5) examined the effects of ingesting a ketone diester on circulating ketone bodies, substrate metabolism and cycling performance under nutritional conditions replicating an elite professional cycling time-trial. Ketone ingestion increased circulating β-hydroxybutyrate and acetoacetate concentrations. Despite optimal nutritional support, the ketone diester was also associated with gut discomfort and an increased perception of effort, leading to an impairment of cycling time-trial performance.
Study 3 (described in chapter 6) manipulated endogenous fat and CHO availability via daily energy intake, to determine whether the metabolic perturbations from a high-fat diet are driven by high-fat or low-CHO availability. Participants consumed five days of a high-fat or highprotein diet (~65% energy intake), while ‘clamping’ CHO consumption to < 20% energy intake. When compared to an isoenergetic high-protein diet, five days’ adaptation to a high-fat diet resulted in greater whole-body rates of fat oxidation during submaximal cycling and decreased skeletal muscle mitochondrial respiration supported by octanoylcarnitine and pyruvate as well as uncoupled respiration at rest. Following one day of a high-CHO diet mitochondrial respiration returned to pre-diet, however whole body rates of substrate oxidation were only partially rescued.
This series of research studies contributes new knowledge to the literature by demonstrating that 1) fat substrates contribute < 20% to energy expenditure during prolonged, high-intensity running, independent of pre-exercise CHO intake 2) ketone diester ingestion impairs cycling time trial performance and is associated with a higher perception of effort, despite optimal nutritional feeding and 3) high dietary fat rather than low-CHO intake contributes to reductions in mitochondrial respiration and increases in whole-body rates of fat oxidation following a high-fat, low-CHO diet. However, this reduction can be partially rescued following one day of a high-CHO diet. This novel information provides evidence that high-fat diets and exogenous ketone drinks are not advantageous for an athletes training and competition due to their detrimental effects on substrate metabolism and skeletal muscle adaptations. Athletes should instead ensure high-CHO availability prior to and during competition to maximise whole-body rates of CHO oxidation rates.
33
Zhang, Xinchen. "Interaction of PEG-ylated Lipid Nanoparticles with Silica Substrates." Thesis, Uppsala universitet, Institutionen för kemi - BMC, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-296349.
Abstract:
In this project, the interaction between polyethylene glycol modified (PEG-ylated) lipid nanoparticles and silica substrates was studied to find out how this interaction was affected by bulk concentration, temperature and the composition of particles. One kind of lipodisk and four kinds of PEG-ylated liposome were prepared from lipid films and characterized by quartz crystal microbalance with dissipation monitoring (QCM-D) instrument mounted with silica sensor. The detailed information of particle-silica interaction could be obtained from the raw data, frequency and dissipation values, and the adsorbed mass surface density calculated from the raw data. Lipodisks could be immobilized on the silica surface. Whether they would be rinsed away by PBS buffer was influenced by both the bulk concentration and temperature. The way of their binding could change and the changing process was affected by temperature. PEG-ylated liposomes could also be immobilized on the silica surface, and they could break and spread to form supported lipid bilayer in certain conditions, for example, the changing of temperature or the using of certain lipids. Supported lipid bilayers were created with high reproducibility in this project, which could be very useful to the future study of transmembrane proteins functions and lipodisk properties.
34
Schafe, Glenn E. "Neuroanatomical substrates of conditioned taste aversion : forebrain-brainstem interactions /." Thesis, Connect to this title online; UW restricted, 1997. http://hdl.handle.net/1773/9068.
35
Aref, Amirreza. "Nanotechnology applied to stem cell-substratum interactions." Thesis, Cranfield University, 2008. http://dspace.lib.cranfield.ac.uk/handle/1826/4448.
Abstract:
The modulation of biological interaction with artificial surfaces is a vital aspect of biomaterials research. Perhaps the most challenging area is transplantation involving the introduction of stem cells into the body with their ability to differentiate; the response of stem cells to implanted biomaterials (or to the host tissue) provides a uniquely sensitive way to explore biocompatibility. An understanding of how to direct specific substratumcellular responses is critical for the development of future biomaterials (e.g., for prosthesis). Attachment and spreading of a cell to and on a substratum are the first part of the process that leads to the ultimate assimilation of the new cell or prosthesis with the host tissue. Together with conventional microscopy, I have exploited a uniquely powerful noninvasive optical technique (Optical Waveguide Lightmode Spectroscopy, OWLS) to quantify cell attachment and spreading of stem cells to artificial biomaterials, and determine how the cell environment (the substratum),the complex liquid medium bathing the cell, and the presence of congeners, influence attachment and spreading. My results highlight that quantitative characterisation of interfacial interactions, including their kinetics leads to uniquely new insight into cell-protein-material interactions. This knowledge will be doubtless be useful in the development of new generations of biomaterials with improved properties designed for specific applications.
36
Markevich, Alexander. "Modification of electronic properties of graphene by interaction with substrates and dopants." Thesis, University of Exeter, 2012. http://hdl.handle.net/10871/10726.
Abstract:
First-principles calculations have been carried out to investigate structural and electronic properties of graphene on SiC and diamond substrates and for a study of doping of fluorographene with various surface adsorbates. New insight is given into the problem of the decoupling of the graphene layers from SiC substrates after epitaxial growth. Mechanisms of hydrogen penetration between graphene and SiC(0001) surface, and properties of hydrogen and fluorine intercalated structures have been studied. Energy barriers for diffusion of atomic and molecular hydrogen through the interface graphene layer with no defects and graphene layers containing Stone-Wales defect or two- and four-vacancy clusters have been calculated. It is argued that diffusion of hydrogen towards the SiC surface occurs through the hollow defects in the interface graphene layer. It is further shown that hydrogen easily migrates between the graphene layer and the SiC substrate and passivates the surface Si bonds, thus causing the graphene layer decoupling. According to the band structure calculations the graphene layer decoupled from the SiC(0001) surface by hydrogen intercalation is undoped, while that obtained by the fluorine intercalation is p-type doped. Further, structure and the electronic properties of single and double layer graphene on H-, OH-, and F- passivated (111) diamond surface have been studied. It is shown that graphene only weakly interacts with the underlying substrates and the linear dispersion of graphene pi-bands is preserved. For graphene on the hydrogenated diamond surfaces the charge transfer results in n-type doping of graphene layers and the splitting of conduction and valence bands in bilayer graphene. For the F- and OH-terminated surfaces, charge transfer and doping of graphene do not occur. Finally, the possibility of doping fluorographene by surface adsorbates have been investigated. The structure and electronic properties of fluorographene with adsorbed K, Li, Au atoms, and F4-TCNQ molecule are described. It is shown that adsorption of K or Li atoms results in electron doping of fluorographene, while Au atoms and F4-TCNQ introduce deep levels inside the band gap. The calculated value of the fluorographene work function is extremely high, 7.3 eV, suggesting that p-type doping is difficult to achieve.
37
Molloy, Claire Ann. "Interaction between oestradiol and the IGF-I signal transduction pathway in breast cancer cells." Thesis, University of Newcastle Upon Tyne, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265214.
38
BERGERAT, AGNES. "Caracterisation des interactions de la methylase dam d'e. Coli avec ses substrats." Paris 6, 1990. http://www.theses.fr/1990PA066406.
Abstract:
La methylase dam (dna-adenine-methylase) d'e. Coli reconnait les sites gatc sur l'adn et transfere un groupement methyl de la s-adenosyl-methionine sur le groupement amino des adenines de ces sites. La methylase dam localise les sequences gatc sur le genome par fixation aleatoire sur l'adn non specifique, puis diffusion jusqu'au site le plus proche. Ce mecanisme de diffusion le long de la double helice est aussi suivi par l'enzyme pour aller d'un site a l'autre. La methylase dam est donc une enzyme processive. La methylase dam presente une affinite particuliere pour chacun des brins d'un site gatc en fonction des sequences adjacentes. Elle interagit donc de facon asymetrique avec le site gatc double brin. L'ado-met joue deux roles aupres de la methylase dam. Elle a un role d'effecteur allosterique. En effet, quand l'enzyme se fixe sur un site gatc en presence d'ado-met elle subit un remaniement de conformation qui renforce son affinite pour ce site et la rend capable de catalyser la reaction de methylation. Ce rearrangement qui pourrait etre l'ajustement induit de l'enzyme autour du site gatc correspond a la formation d'un complexe ternaire actif. Elle a aussi un deuxieme role, catalytique en tant que donneur de groupements methyl. Pour remplir chacune de ces fonctions l'ado-met se fixe a l'enzyme sur deux sites distincts avec deux affinites differentes
39
ESSIA, NGANG JEAN-JUSTIN. "Interaction s. Cerevisiae l. Casei en fermentation alcoolique de substrats de sucrerie." Amiens, 1991. http://www.theses.fr/1991AMIES006.
Abstract:
La fermentation alcoolique industrielle de sous-produits de sucrerie, se deroule en conditions non steriles, ce qui entraine le developpement de bacteries et notamment de germes lactiques. Cette etude montre les perturbations induites par l. Casei sur s. Cerevisiae. L'acide lactique, actif par sa forme non dissociee, modifie les besoins en maintenance ainsi que le stock d'acides gras de s. Cerevisiae. Toutefois, la part du metabolite dans l'inhibition est faible et la presence de la cellule bacterienne est responsable de la reponse des levures. S. Cerevisiae favorise la croissance de l. Casei par son activite invertasique mais elle est d'autant moins inhibee que la taille de l'inoculum est importante. Un recyclage controle de la biomasse permet de rester en deca d'une population critique en contaminants
40
Fransson, Linda. "Enzyme substrate solvent interactions : a case study on serine hydrolases." Doctoral thesis, KTH, Biokemi, 2008. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-4867.
Abstract:
Reaction rates and selectivities were measured for transacylation of fatty acid esters in solvents catalysed by Candida antarctica lipase B and by cutinase from Humicola insolens. With these enzymes classical water-based enzymology can be expanded to many different solvents allowing large variations in interaction energies between the enzymes, the substrates and the surrounding. Further ,hydrolysis reactions catalysed by Bacillus subtilis esterase 2 were investigated. Thermodynamics analyses revealed that the enzyme contribution to reaction rate acceleration compared to acid catalysis was purely entropic. On the other hand, studies of differences in activation entropy and enthalpy between enantiomers and between homologous esters showed that high substrate specificity was favoured by enthalpic stabilisation. Solvent was found to have a profound effect on enzyme catalysis, affecting both reaction rate and selectivity. Differences in substrate solubility will impact enzyme specificity since substrate binding is an equilibrium between enzyme-bound substrate and substrate in free solution. In addition, solven tmolecules were found to act as enzyme inhibitors, showing both competitive and non-competitive behaviour. In several homologous data series enthalpy-entropy compensation relationships were encountered. A possible extrathermodynamic relationship between enthalpy and entropy can easily be lost under co-varying errors propagated from the experiments. From the data in this thesis, one instance was found of a real enthalpy-entropy compensation that could be distinguished from statistical errors, while other examples could not be verified.QC 20100722
41
Meyer, Andrew James. "A calorimetric study of host-guest and protein-substrate interactions." Thesis, University of Leicester, 1997. http://hdl.handle.net/2381/30009.
Abstract:
This thesis describes a study of molecular recognition processes involving - and - cyclodextrins, and the enzymes chloramphenicol acetyltransferase and DNA gyrase.;An isothermal titration calorimeter of high sensitivity was used to investigate the binding of a number of relatively small guest molecules with much larger host molecules. The calorimetric technique allows the direct determination of the apparent binding enthalpy bindH0', the evaluation of the apparent association constant Ka' and hence evaluation of the apparent Gibbs energy and entropy of binding, bindG0' and TbindS0'.;The energetics of association for p-nitrophenol and 1-adamantane carboxylic acid in neutral and anionic forms, binding to - and - cyclodextrins were measured, and the results compared with those from previous studies. Binding of p-fluoronitrobenzene, p-nitrobenzyl alcohol and the antibiotic chloramphenicol to the cyclodextrins was also studied and the thermodynamic binding parameters identified. The results showed that where there is a close fit of the guest molecule inside the cyclodextrin cavity, the binding is enthalpy driven. Where there is a loose fit the binding is entropically favourable, but is still enthalpy dominated.;Binding of hydrophobic derivatives of the antibiotic chloramphenicol (CM), to the enzyme chloramphenicol acetyltransferase (CAT), was investigated. Six CM analogues were prepared with substitutions made at the 3-hydroxy group of CM, and enthalpies of binding to CAT determined by calorimetry. Comparison of calorimetric data with Gibbs energies of binding determined previously, showed that the binding is enthalpy driven.;Binding of coumarin drugs to the 43-kDa and 24-kDa protein fragments of the enzyme DNA gyrase was studied using titration calorimetry. The energetics of binding obtained from the calorimetric technique were compared with those obtained from rapid gel filtration (spin-column) studies. Titration calorimetry shows that the Ka' for coumarins binding to both proteins is of the order of 108 mol-1dm3 and that the binding is enthalpy driven.
42
Yap, Jessica. "Identification of Plasmodium falciparum protein kinase substrates and interacting proteins." Honors in the Major Thesis, University of Central Florida, 2012. http://digital.library.ucf.edu/cdm/ref/collection/ETH/id/644.
Abstract:
Characterization of PfPKA and PfPK5 substrates, as well as the proteins they interact with, will help us to develop innovative therapies targeting binding sites.; Malaria is a devastating disease that results in almost one million deaths annually. Most of the victims are children under the age of five in Sub-Saharan Africa. Malaria parasite strains throughout developing countries are continually building resistance to available drugs. Current therapies such as mefloquine, chloroquine, as well as artemisinin are becoming less effective, and this underscores the urgency for therapeutics directed against novel drug targets. In order to identify new drug targets, the molecular biology of the malaria parasite Plasmodium needs to be elucidated. Plasmodium exhibits a unique cell cycle in which it undergoes multiple rounds of DNA synthesis and mitosis without cytokinesis. Thus, cell cycle regulatory proteins are likely to be promising pathogen-specific drug targets. It is expected that fluctuating activity of key proteins, such as protein kinases, play an essential role in regulating the noncanonical life cycle of Plasmodium. Consequently, malarial kinases are a prime target for therapy. One way to better understand the role of malarial kinases in Plasmodium cell cycle regulation is to identify putative protein kinase substrates and interacting proteins. Two malarial kinases that have been implicated in regulating malaria parasite cell cycle stages were investigated in this study: P. falciparum CDK-like Protein Kinase 5 (PfPK5) and cAMP-Dependent Protein Kinase A (PfPKA). A transgenic P. falciparum line was created for the expression of epitope-tagged PfPK5 for pull-down analysis. Phospho-substrate antibodies were used to identify physiological substrates of both PfPK5 and PfPKA. Immunoblotting with these antibodies identified several potential substrates. Identities of the PfPKA physiological substrates were determined from the global P. falciparum phosphoproteome dataset that has recently been generated in our laboratory.B.S.
Bachelors
Burnett School of Biomedical Sciences
Molecular and Microbiology
43
Öhlund, Thomas. "Metal Films for Printed Electronics : Ink-substrate Interactions and Sintering." Doctoral thesis, Mittuniversitetet, Avdelningen för naturvetenskap, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:miun:diva-23420.
Abstract:
A new manufacturing paradigm may lower the cost and environmental impact of existing products, as well as enable completely new products. Large scale, roll-to-roll manufacturing of flexible electronics and other functionality has great potential. However, a commercial breakthrough depends on a lower consumption of materials and energy compared with competing alternatives, and that sufficiently high performance and reliability of the products can be maintained. The substrate constitutes a large part of the product, and therefore its cost and environmental sustainability are important. Electrically conducting thin films are required in many functional devices and applications. In demanding applications, metal films offer the highest conductivity. In this thesis, paper substrates of various type and construction were characterized, and the characteristics were related to the performance of inkjet-printed metal patterns. Fast absorption of the ink carrier was beneficial for well-defined pattern geometry, as well as high conductivity. Surface roughness with topography variations of sufficiently large amplitude and frequency, was detrimental to the pattern definition and conductivity. Porosity was another important factor, where the characteristic pore size was much more important than the total pore volume. Apparent surface energy was important for non-absorbing substrates, but of limited importance for coatings with a high absorption rate. Applying thin polymer–based coatings on flexible non-porous films to provide a mechanism for ink solvent removal, improved the pattern definition significantly. Inkjet-printing of a ZnO-dispersion on uncoated paper provided a thin spot-coating, allowing conductivity of silver nanoparticle films. Conductive nanoparticle films could not form directly on the uncoated paper. The resulting performance of printed metal patterns was highly dependent on a well adapted sintering methodology. Several sintering methods were examined in this thesis, including conventional oven sintering, electrical sintering, microwave sintering, chemical sintering and intense pulsed light sintering. Specially designed coated papers with modified chemical and physical properties, were utilized for chemical low-temperature sintering of silver nanoparticle inks. For intense pulsed light sintering and material conversion of patterns, custom equipment was designed and built. Using the equipment, inkjet-printed copper oxide patterns were processed into highly conducting copper patterns. Custom-designed papers with mesoporous coatings and porous precoatings improved the reliablility and performance of the reduction and sintering process. The thesis aims to clarify how ink-substrate interactions and sintering methodology affect the performance and reliability of inkjet-printed nanoparticle patterns on flexible substrates. This improves the selection, adaptation, design and manufacturing of suitable substrates for inkjet-printed high conductivity patterns, such as circuit boards or RFID antennas.
44
Al-Hamdani, Y. S. "Theoretical approach towards accurate molecular interactions with low dimensional substrates." Thesis, University College London (University of London), 2016. http://discovery.ucl.ac.uk/1531128/.
45
Lack, Barbara Anne. "Metal interactions with neural substrates and their role in neurodegeneration." Thesis, Rhodes University, 2003. http://hdl.handle.net/10962/d1005709.
Abstract:
"Life" may be characterized as a controlled stationary flow equilibrium, maintained by energy consuming chemical reactions. The physiological functioning of these life systems include at least 28 of the elements isolated on the periodic table thus far, most of which are metals. However, as with Paracelsus Principle: "The dose makes the poison", there exists a definite link between metal levels, essential and toxic, and the onset of neurodegenerative diseases. The economic costs of brain dysfunction are enormous, but this pales in comparison to the staggering emotional toll on the victims themselves and their families. In an attempt to improve the understanding of the causes of neurodegeneration, this study focuses on one potential aspect: the possible link between metals and neurotransmitter homeostasis utilising a variety of electronanalytical techniques. Adsorptive cathodic stripping voltammetry was employed to investigate the binding affinities and complex formation of melatonin and its precursor serotonin with calcium, potassium, sodium, lithium and aluminium. The results showed that all the metals studied formed complexes with both pineal indoleamines. However, the stability and affmity of the ligands toward the various metals varied greatly. The study suggests a further role for melatonin, that of metalloregulator and possible metal detoxifier in the brain, the in vivo studies which followed will further substantiate this notion. This research additionally focused on the cholinergic system, in particular acetylcholine complex formation studies with mercury, lead, cadmium, copper and zinc using the adsorptive cathodic stripping voltammetry method. The formation and characterisation of a solid mercury-acetylcholine complex lent further strength to the in situ electrochemical complex formation observed. The results showed the preference of acetylcholine for environmentally toxic heavy metals (such as Cd²⁺) over those divalent cations that occur naturally in the body. The possible metalloregulatory role melatonin played in the three brain regIOns: cerebellum, cortex and corpus striatum of male Wistar rats was studied as an in vivo extension of the earlier in vitro studies. Anodic stripping voltammetry was employed to detect metal levels present. The results showed that daily injections of melatonin was responsible for significantly decreasing copper(I), cadmium(II) and lead(II) levels in various regions of the rat brain of those animals that had undergone a pinealectomy in comparison to the saline injected group having undergone the same treatment. Histological and electrochemical stripping techniques were applied to investigate the implications of high A1³⁺ levels in the brain regions, particularly the hippocampus. Melatonin showed signs of promise in indirect symptom alleviation and by significantly decreasing A1³⁺ levels in rats that had been dosed with melatonin prior to A1³⁺ treatments in comparison with the control groups. Finally a preliminary study outlining a method for the production of a calcium selective microelectrode was undertaken. Further work is still needed to optimise the microelectrode production as well as its possible applications. However, whilst the overall conclusions of this entire multidisciplinary study may indeed only be in effect one piece of a very large puzzle on neurodegenerative diseases, this piece will no doubt serve as a building block for further ideas and work in this field.
46
Russ, Jennifer Lynn. "Studies of Solution Paramagnetic-Substrate Nuclear and Electron Intermolecular Interactions." Diss., Virginia Tech, 2006. http://hdl.handle.net/10919/27050.
Abstract:
Advanced nuclear and electron magnetic resonance techniques (i.e. nuclear magnetic resonance (NMR), dynamic nuclear polarization (DNP), and magnetic resonance imaging (MRI)) were used to study the attitude and dynamics of TEMPO (2,2,6,6-tetramethylpiperidinyloxy)-substrate systems and the relaxivity properties of water-soluble trimetallic nitride template functionalized endohedral metallofullerenes (TNT-fMF). The attitude and average distance of interaction for each TEMPO-substrate system was determined from comparing density functional theory (DFT) calculation results with experimental hyperfine coupling constants leading to an improved understanding of solution dynamics. The short-lived solvent-solute interactions of the TEMPO-substrate molecules, such as transient complex formation, are governed by weak hydrogen-bonding interactions. The collisions in solution were explained by determining the favored orientations of the two molecules interacting using calculated relative energy minima and reproducibility of the experimental results by the calculated coupling constants.
Water-soluble TNT-fMFs are studied as candidates for the next generation MRI contrast agents as diagnostic agents and also as possible therapeutic agents to kill cancer cells and decrease tumors. The TNT-fMFs are being studied as part of a multi-modal platform dependent upon which metal atoms are encapsulated inside: Gd â MRI contrast agent (diagnostic), Lu and Ho â radio labeled for use as a therapeutic agent, Tb â fluorescence, and Lu â x-ray contrast. The current commercial MRI contrast agent, OmniscanTM, contains one gadolinium atom; however, the metal is complexed to, not encapsulated in, the molecule. TNT-fMFs fully encapsulate three metal atoms to ensure the patient does not run the risk of metal poisoning. The r1 and r2 relaxivities of TNT-fMFs containing either Gd, Lu, Ho, or Sc metals were measured at 0.35T. The data for the Gd containing TNT-fMFs indicated the metallofullerene has significantly higher relaxivities than OmniscanTM, and can be the next generation MRI contrast agent. The Ho containing species has a high r2/r1 ratio compared to the other samples showing it is a potential T2 agent, and has therapeutic capabilities.Ph. D.
47
Bach, Kristensen Jan. "Enzymatic hydrolysis of lignocellulose : substrate interactions and high solids loadings." Forest & Landscape, 2008. http://www.sl.kvl.dk/upload/nr_42_phd_jan__web.pdf.
48
Panse, Vikram G. "Interaction Of Chaperone SecB With Protein Substrates: A Biophysical Study." Thesis, Indian Institute of Science, 2000. https://etd.iisc.ac.in/handle/2005/242.
Abstract:
In the cell, as in in vitro, the final conformation of a protein is determined by it's amino acid sequence (1). Some isolated proteins can be denatured and refolded in vitro in absence of extrinsic factors. However, in order to fold in the cell, the newly synthesized polypeptide chain has to negotiate an environment far more complex than that faced by the unfolded chain in vitro. Cells have evolved proteins called “chaperones” to assist folding and assembly of polypeptides (2). Thus, the linear sequence of a protein not only contains information that specifies the final three-dimensional functional form, but also recognition motifs, which can be recognized by the cellular folding machinery.
The work reported in this thesis is aimed at understanding some aspects of recognition of target substrates by the cytosolic chaperone, SecB, which forms part of the protein translocation machinery in E. coli. The sec pathway is involved in both translocation of precursor proteins across and the insertion of integral membrane proteins into the cytoplasmic membrane (3).
Chapter one discusses some general aspects of protein folding and briefly describes chaperone systems, which have been extensively characterized in literature.
Chapter two discusses the effect of chaperone SecB on the refolding pathway of a model substrate protein barstar, whose folding pathway has been extensively characterized (4,5). The effect of SecB on the refolding kinetics of the small protein barstar (wild type) and
fluorescein labeled C82A (single Cys mutant) in 1 M guanidine hydrochloride at pH 7.0 at 25 °C has been investigated using fluorescence spectroscopy. We show that SecB does not bind either the native or the unfolded states of barstar but binds to late near-native intermediate (s) along the folding pathway. ESR studies and fluorescence anisotropy measurements show that SecB forms stable complexes with the near-native intermediate (s). For barstar, polypeptide collapse and formation of a hydrophobic surface are required for binding to SecB. Steady state polarization measurements indicated the presence of stable complexes of barstar bound to SecB. Studies on the spin labeled C82A show an immobilization of the spin label adduct at the 40th position of barstar, suggesting that the binding of SecB to barstar occurs in that region. SecB does not change the apparent rate constant of barstar refolding. The kinetic data for SecB binding to barstar are not consistent with simple kinetic partitioning models (6).
Chapter three discusses the energetics of substrate:SecB interactions using the following model protein substrates: unfolded RNase A, BPTI, partially folded disulfide intermediates of alpha-lactalbumin,. The thermodynamics of binding of unfolded polypeptides to the chaperone SecB were investigated in vitro by isothermal titration calorimetry and fluorescence spectroscopy. The heat capacity changes observed on binding the reduced and carboxamidomethylated forms of alpha-lactalbumin, BPTI, and RNase A were found to be -0.10, -0.29 and -0.41 kcal mol-1 K-1 respectively and suggest that between 7 and 29 residues are buried upon substrate binding to SecB. In all cases binding occurs with a stoichiometry of one polypeptide chain per monomer of SecB. The data are consistent with a model where SecB binds substrate molecules at an exposed hydrophobic cleft (7).
Chapter four discusses the thermodynamics of unfolding to gain insights into the mechanism of assembly and stability of the tetrameric structure. The thermodynamics of unfolding of SecB was studied as a function of protein concentration, by using high sensitivity-differential scanning calorimetry and spectroscopic methods. The thermal unfolding of tetrameric SecB is reversible and can be well described as a two-state transition in which the folded tetramer is converted directly to unfolded monomers. The
value of ACP obtained was 10.7 ± 0.7 kcal mol-1 K-1, which is amongst the highest measured for a multimeric protein. At 298 K, pH 7.4. the AG°U for the SecB tetramer is 27.9 ± 2 kcal mol-1. Denaturant mediated unfolding of SecB was found to be irreversible. The reactivity of the 4 solvent exposed free thiols in tetrameric SecB is salt dependent. The kinetics of reactivity suggests that these four Cysteines are in close proximity to each other and that these residues on each monomer are in chemically identical environments. The thermodynamic data suggest that SecB is a stable, well folded and tightly packed tetramer and that substrate binding occurs at a surface site rather than at an interior cavity (8).
Chapter five discusses the bound state conformation of a model protein substrate of SecB, bovine pancreatic trypsin inhibitor (BPTI), as well as the conformation of SecB itself by using proximity relationships based on site-directed spin-labeling and pyrene fluorescence methods. BPTI is a 58 residue protein and contains 3 disulfide groups between residues 5 and 55, 14 and 38, and 30 and 51. Single disulfide mutants of BPTI were reduced and the free cysteines were labeled with either thiol-specific spin labels or pyrene maleimide. The relative proximity of labeled residues was studied using either electron spin resonance spectroscopy or fluorescence spectroscopy. The data suggest that SecB binds a collapsed coil of reduced unfolded BPTI, which then undergoes a structural rearrangement to a more extended state upon binding to SecB. Binding occurs at multiple sites on the substrate and the binding site on each SecB monomer accommodates less than 21 substrate residues. In addition, we have labeled four, solvent accessible cysteine residues in the SecB tetramer and have investigated their relative spatial arrangement in the presence and absence of the substrate protein. The ESR data suggest that these cysteine residues are in close proximity when no substrate protein is bound, but move away from each other when SecB binds substrate. This is the first direct evidence of a conformational change in SecB upon binding of a substrate protein.
Chapter six discusses the mechanism of dissaggregation of a model peptide aggregate by chaperone SecB. The Hspl04, Hsp70 and Hsp40 chaperone system are capable of dissociating aggregated state(s) of substrate proteins, though little is known of the
mechanism of the process. The interaction of the B chain of insulin with chaperone SecB was investigated using light scattering, pyrene excimer fluorescence and electron spin resonance spectroscopy. We show that SecB prevents aggregation of the B chain of insulin. We show that SecB is capable of dissociating soluble B chain aggregate as monitored by pyrene fluorescence spectroscopy. The kinetics of dissociation of the B chain aggregate by SecB has also been investigated to understand the mechanism of dissociation. The data suggests that SecB does not act as a catalyst in dissociation of the aggregate to individual B chains, rather it binds the small population of free B chains with high affinity, thereby shifting the equilibrium from the ensemble of the aggregate towards the individual B chains. Thus SecB can rescue aggregated, partially folded /misfolded states of target proteins by a thermodynamic coupling mechanism when the free energy of binding to SecB is greater than the stability of the aggregate. Pyrene excimer fluorescence and ESR methods have been used to gain insights on the bound state conformation of the B chain to chaperone SecB. The data suggests that the B chain is bound to SecB in a flexible extended state in a hydrophobic cleft on SecB and that the binding site accommodates approximately 10 residues of substrate (9).
49
Panse, Vikram G. "Interaction Of Chaperone SecB With Protein Substrates: A Biophysical Study." Thesis, Indian Institute of Science, 2000. http://hdl.handle.net/2005/242.
Abstract:
In the cell, as in in vitro, the final conformation of a protein is determined by it's amino acid sequence (1). Some isolated proteins can be denatured and refolded in vitro in absence of extrinsic factors. However, in order to fold in the cell, the newly synthesized polypeptide chain has to negotiate an environment far more complex than that faced by the unfolded chain in vitro. Cells have evolved proteins called “chaperones” to assist folding and assembly of polypeptides (2). Thus, the linear sequence of a protein not only contains information that specifies the final three-dimensional functional form, but also recognition motifs, which can be recognized by the cellular folding machinery.
The work reported in this thesis is aimed at understanding some aspects of recognition of target substrates by the cytosolic chaperone, SecB, which forms part of the protein translocation machinery in E. coli. The sec pathway is involved in both translocation of precursor proteins across and the insertion of integral membrane proteins into the cytoplasmic membrane (3).
Chapter one discusses some general aspects of protein folding and briefly describes chaperone systems, which have been extensively characterized in literature.
Chapter two discusses the effect of chaperone SecB on the refolding pathway of a model substrate protein barstar, whose folding pathway has been extensively characterized (4,5). The effect of SecB on the refolding kinetics of the small protein barstar (wild type) and
fluorescein labeled C82A (single Cys mutant) in 1 M guanidine hydrochloride at pH 7.0 at 25 °C has been investigated using fluorescence spectroscopy. We show that SecB does not bind either the native or the unfolded states of barstar but binds to late near-native intermediate (s) along the folding pathway. ESR studies and fluorescence anisotropy measurements show that SecB forms stable complexes with the near-native intermediate (s). For barstar, polypeptide collapse and formation of a hydrophobic surface are required for binding to SecB. Steady state polarization measurements indicated the presence of stable complexes of barstar bound to SecB. Studies on the spin labeled C82A show an immobilization of the spin label adduct at the 40th position of barstar, suggesting that the binding of SecB to barstar occurs in that region. SecB does not change the apparent rate constant of barstar refolding. The kinetic data for SecB binding to barstar are not consistent with simple kinetic partitioning models (6).
Chapter three discusses the energetics of substrate:SecB interactions using the following model protein substrates: unfolded RNase A, BPTI, partially folded disulfide intermediates of alpha-lactalbumin,. The thermodynamics of binding of unfolded polypeptides to the chaperone SecB were investigated in vitro by isothermal titration calorimetry and fluorescence spectroscopy. The heat capacity changes observed on binding the reduced and carboxamidomethylated forms of alpha-lactalbumin, BPTI, and RNase A were found to be -0.10, -0.29 and -0.41 kcal mol-1 K-1 respectively and suggest that between 7 and 29 residues are buried upon substrate binding to SecB. In all cases binding occurs with a stoichiometry of one polypeptide chain per monomer of SecB. The data are consistent with a model where SecB binds substrate molecules at an exposed hydrophobic cleft (7).
Chapter four discusses the thermodynamics of unfolding to gain insights into the mechanism of assembly and stability of the tetrameric structure. The thermodynamics of unfolding of SecB was studied as a function of protein concentration, by using high sensitivity-differential scanning calorimetry and spectroscopic methods. The thermal unfolding of tetrameric SecB is reversible and can be well described as a two-state transition in which the folded tetramer is converted directly to unfolded monomers. The
value of ACP obtained was 10.7 ± 0.7 kcal mol-1 K-1, which is amongst the highest measured for a multimeric protein. At 298 K, pH 7.4. the AG°U for the SecB tetramer is 27.9 ± 2 kcal mol-1. Denaturant mediated unfolding of SecB was found to be irreversible. The reactivity of the 4 solvent exposed free thiols in tetrameric SecB is salt dependent. The kinetics of reactivity suggests that these four Cysteines are in close proximity to each other and that these residues on each monomer are in chemically identical environments. The thermodynamic data suggest that SecB is a stable, well folded and tightly packed tetramer and that substrate binding occurs at a surface site rather than at an interior cavity (8).
Chapter five discusses the bound state conformation of a model protein substrate of SecB, bovine pancreatic trypsin inhibitor (BPTI), as well as the conformation of SecB itself by using proximity relationships based on site-directed spin-labeling and pyrene fluorescence methods. BPTI is a 58 residue protein and contains 3 disulfide groups between residues 5 and 55, 14 and 38, and 30 and 51. Single disulfide mutants of BPTI were reduced and the free cysteines were labeled with either thiol-specific spin labels or pyrene maleimide. The relative proximity of labeled residues was studied using either electron spin resonance spectroscopy or fluorescence spectroscopy. The data suggest that SecB binds a collapsed coil of reduced unfolded BPTI, which then undergoes a structural rearrangement to a more extended state upon binding to SecB. Binding occurs at multiple sites on the substrate and the binding site on each SecB monomer accommodates less than 21 substrate residues. In addition, we have labeled four, solvent accessible cysteine residues in the SecB tetramer and have investigated their relative spatial arrangement in the presence and absence of the substrate protein. The ESR data suggest that these cysteine residues are in close proximity when no substrate protein is bound, but move away from each other when SecB binds substrate. This is the first direct evidence of a conformational change in SecB upon binding of a substrate protein.
Chapter six discusses the mechanism of dissaggregation of a model peptide aggregate by chaperone SecB. The Hspl04, Hsp70 and Hsp40 chaperone system are capable of dissociating aggregated state(s) of substrate proteins, though little is known of the
mechanism of the process. The interaction of the B chain of insulin with chaperone SecB was investigated using light scattering, pyrene excimer fluorescence and electron spin resonance spectroscopy. We show that SecB prevents aggregation of the B chain of insulin. We show that SecB is capable of dissociating soluble B chain aggregate as monitored by pyrene fluorescence spectroscopy. The kinetics of dissociation of the B chain aggregate by SecB has also been investigated to understand the mechanism of dissociation. The data suggests that SecB does not act as a catalyst in dissociation of the aggregate to individual B chains, rather it binds the small population of free B chains with high affinity, thereby shifting the equilibrium from the ensemble of the aggregate towards the individual B chains. Thus SecB can rescue aggregated, partially folded /misfolded states of target proteins by a thermodynamic coupling mechanism when the free energy of binding to SecB is greater than the stability of the aggregate. Pyrene excimer fluorescence and ESR methods have been used to gain insights on the bound state conformation of the B chain to chaperone SecB. The data suggests that the B chain is bound to SecB in a flexible extended state in a hydrophobic cleft on SecB and that the binding site accommodates approximately 10 residues of substrate (9).
50
Ben, Arfi Rim. "Adsorption, interaction et conformation de molécules modèles d’agent de couplage sur substrats métalliques." Mulhouse, 2007. http://www.theses.fr/2007MULH0886.
Abstract:
Le contexte de l’étude concerne les systèmes composites et notamment l’utilisation d’agents de couplage mis en œuvre pour assurer la liaison entre un substrat métallique et une matrice élastomère. Dans le cadre de cette étude, nous nous sommes intéressés plus particulièrement au cas de l’adsorption du 1-hexanedecanethiol (HS-(CH2)15-CH3) sur différents substrats, représentatifs de l’application industrielle. Par ailleurs, une étude complète a également été menée sur l’adsorption du 1-hexandécylamine (H2N-(CH2)15-CH3). Ces molécules ne diffèrent que par leur fonctionnalité terminale, la longueur de la chaîne alkyle étant identique. Elles permettront ainsi de mettre en évidence l’influence de la réactivité interfaciale sur l’adsorption. Afin de faciliter la caractérisation ultérieure des dépôts, l’étude a été réalisée sur des surfaces planes modèles. Cette étude a clairement permis de mettre en évidence l’adsorption irréversible et homogène de ces moélcules sur les différents substrats et de déterminer précisément l’interaction et l’organisation de ces molécules sondes lors de leur adsorption en surface. L’ensemble des résultats présentés dans cette étude démontrent tout l’intérêt des approches multi-techniques et multi-échelles dans la caractérisation des couches ultra-minces organiquesThe understanding of mechanisms governing the growth, the structure and the conformation of coupling agents onto different metal substrates is determinant for an optimal use in any application. A variety of analytical techniques were used to characterize the different substrates : wettability, ellipsometry, atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS)and the polarization infrared reflection absorption spectroscopy (PM-IRRAS). Observations suggest that the structure of organic films is controlled by varying the concentration of the solution and the assembly time. The packing density, the organization and the conformation of the molecular chain depend on the nature of the metal substrate and on the roughness of the surface. All results presented in this work demonstrate the interest of multi-techniques and multi-scales approach in the characterization of ultra-thin organic films