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Oberirdische Speicher in Segmentbauweise für Wärmeversorgungssysteme – OBSERW: Abschlussbericht zum VerbundvorhabenUrbaneck, Thorsten, Findeisen, Fabian, Mücke, Jan Markus, Lang, Stephan, Gensbaur, Markus, Bestenlehner, Dominik, Drück, Harald, Beyer, Robert, Pieper, Konrad 15 November 2018 (has links)
Im Projekt wurde eine alternative Speicherkonstruktion im Bereich von 500 bis 6000 m3 für den Betrieb in Solar- und Fernwärmesystemen entwickelt. Ausgangspunkt bilden große Kaltwasserspeicher in Segmentbauweise. Die Bautechnologie bietet ein signifikantes Kostenreduktionspotenzial gegenüber
geschweißten Flachbodentanks, konnte bisher aber nicht auf Wärmespeicher übertragen werden. Aufgrund der dünnwandigen Bauweise und der Projektziele musste eine Überarbeitung des Wandaufbaus, der Einbauten und der Peripherie erfolgen.
Dieser Bericht liefert eine Beschreibung des Speicher-Systems und die Ergebnisse des Verbundvorhabens. Die Funktionsfähigkeit wurde mit einem dreistufigen Verfahren nachgewiesen. Das geplante Vorgehen mit Laborversuchen im kleinen Maßstab bis zum Test mit einem Demonstrator im Realmaßstab (100 m3) war notwendig und zielführend. Die Bearbeitung der Hauptaufgaben (z. B. Materialuntersuchungen, Konstruktion, Betrieb) erfolgte vernetzt durch die beteiligten
Forschungsinstitutionen. Das grundlegende Potenzial für eine spätere Anwendung in solaren Nahwärmesystemen oder Sekundärnetzgebieten der klassischen Fernwärme sind gegeben. Vor allem im Bereich der Beladung und im Wandaufbau konnten große Verbesserungen erzielt werden. Weitere Optimierungen und die Umsetzung mit größeren Speichern stehen noch aus. / In the project, an alternative construction for thermal energy stores in the range of 500 to 6000 m3 was developed for operation in solar and district heating systems. Large cold water storage tanks in segmental construction are the starting point. Their construction technology offers a significant potential for cost reduction compared to welded flat-bottom tanks, but could so far not be transferred to hot water storage tanks. Due to the new design and the project objectives, the wall structure, the internals and the periphery had to be completely revised.
This report provides a description of the storage system and the results of the joint project. The functionality was proven with a three-stage procedure. The planned procedure with laboratory tests on a small scale up to the test with a demonstrator on a real scale (100 m3) was necessary and
purposeful. The main tasks (e.g. material testing, design, operation) were carried out by the participating research institutions in a network. The basic potential for a later application in solar local heating systems or secondary network areas of conventional district heating is given. Significant improvements were realized, especially in regard of the charging system and the wall construction.
However, further optimizations and the transfer to larger storage tanks is still pending.
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Numerische Untersuchung der Rayleigh-Bénard-Konvektion in einem Flüssigmetall unter dem Einfluss einer zeitlich modulierten gezeitenartigen KraftRöhrborn, Sebastian 01 September 2023 (has links)
In der vorliegenden Arbeit konnte gezeigt werden, dass die numerischen simulationen einer freien Rayleigh-Bénard-Konvektion und einer rein elektromagnetisch angetriebenen gezeiten-artigen Strömung in einem stehenden zylindrischen Volumen mit einem Seitenverhältnis Г = D/H = 1 und seitlich angelegten Magnetspulen eine gute Übereinstimmung mit entspre-chenden Experimenten aufweisen. Kombiniert man beide Mechanismen und moduliert die Lorentzkraft, so zeigen sich in den Frequenzspektren der Helizität in zwei Halbräumen des Volumens deutliche Maxima an der Modulationsfrequenz. Eine solche Helizitätssynchronisierung durch Gezeitenkräfte wird derzeit als mögliche Erklärung für die hohe Regularität des Sonnendynamos diskutiert. Des Weiteren wird die in freier Konvektion auftretende langsame azimutale Wanderung der Konvektionszelle unterdrückt. Der Schwingungswinkel der azimutalen Schwappbewegung nimmt dabei ab und die in der Strömung dominante Frequenz erhöht sich. Die durch die zwei unterschiedlichen Antriebsmechanismen erzeugten Strömungsstrukturen bleiben in der Strömung eigenständig erhalten und treten in gegenseitige Interaktion.:1. Einleitung
2. Grundlagen
2.1. Rayleigh-Bénard-Konvektion
2.2. MHD - Magnetohydrodynamik
2.3. Wichtige Aspekte des numerischen Modells
3. Modellerstellung
3.1. Geometrie
3.2. Numerisches Modell
3.2.1. Elektromagnetisches Modell in Opera
3.2.2. Modell der Strömungsberechnung in OpenFOAM
4. Ergebnisse
4.1. Ergebnisse der freien Rayleigh-Bénard-Konvektion
4.2. Ergebnisse der nichtmodulierten elektromagnetischen Strömungsanregung ohne Temperaturgradient
4.3. Ergebnisse der zeitmodulierten elektromagnetischen Strömungsanregung ohne Temperaturgradient
4.4. Ergebnisse der elektromagnetisch beeinflussten Rayleigh-Bénard-Konvektion
4.4.1. Auswirkung der elektromagnetischen Beeinflussung auf die Strömungsstruktur
4.4.2. Vergleich ausgewählter Ergebnisse der numerischen Untersuchung und des Experimentes
4.4.3. Auswirkung der elektromagnetischen Beeinflussung auf die Helizität
5. Zusammenfassung und Fazit
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Modelluntersuchungen zu Filtrationsmechanismen in keramischen SchaumfilternHeuzeroth, Fabian 21 March 2017 (has links) (PDF)
Die eingereichte Arbeit ist ein Beitrag, um das Verständnis der Filtration von metallischen Schmelzen für hochwertige Gussprodukte zu erhöhen. Anhand theoretischer und experimenteller Betrachtungen wird besonders die Benetzung zwischen der kontinuierlichen und der dispersen Phase als Einflussfaktor auf Tiefenfiltrationsprozesse untersucht. Rasterkraftmikroskopische Messungen verdeutlichen die Bedeutung der Benetzung auf die Adhäsionsenergie zwischen zwei Feststoffoberflächen. Durch ein entwickeltes Modellsystem konnte bewiesen werden, dass mit steigendem Kontaktwinkel zwischen disperser und kontinuierlicher Phase die Filtrationseffizienz signifikant ansteigt. Aufgrund der wirkenden Haftmechanismen im getesteten Benetzungssystem können sich dendritische Abscheidestrukturen ausbilden, die zur Redispergierung neigen. Ergänzt um eine numerische Simulation am Einzelkollektor, konnte ein Abscheidemodell entwickelt werden. Dieses berücksichtigt erstmals reale Haftenergieverteilungen, simulierte Aufprallenergieverteilungen und die hydrodynamische Hemmung als relevante Parameter für die Tiefenfiltration.
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Qualidade da madeira de espécies do gênero Acacia plantadas no Brasil / Wood quality of species of the genus Acacia planted in BrazilAlencar, Gláucia Soares Barbosa de 09 June 2009 (has links)
O gênero Acacia apresenta espécies florestais de rápido crescimento, com rotações curtas (7 a 9 anos) e de grande interesse comercial para produção de polpa celulósica, painéis de madeira e energia. No Brasil a utilização de espécies do gênero Acacia na produção de celulose de fibra curta ainda é bastante restrita e são poucos os estudos que demonstram a potencialidade do uso dessa matéria-prima para a obtenção de polpa celulósica com as propriedades alcançadas pelas espécies do gênero Eucalyptus. Este trabalho teve como objetivo analisar o potencial de algumas espécies do gênero Acacia (A.aulacocarpa, A. auriculiformis, A. crassicarpa, A. mangium) plantados no Brasil, para a produção de polpa celulósica Kraft, considerando os aspectos relacionados à qualidade da madeira e da polpa, o consumo de reagentes químicos na polpação e também os aspectos relacionados ao rendimento e a capacidade de produção de polpa celulósica; adicionalmente buscou-se fornecer informações estratégicas para o setor celulósico nacional com relação as possíveis ameaças e as oportunidades apresentadas pelos materiais não utilizados no Brasil. Para realização deste trabalho foram utilizados 11 materiais genéticos distintos e cada um dos materiais genéticos foi considerado um tratamento e todos com 5 anos de idade. As características avaliadas foram: densidade básica, dimensões das fibras, composição química e parâmetros da polpação kraft (rendimento bruto, rendimento depurado, teor de rejeitos, número kappa e relações). Para este último objetivo, as madeiras de cada uma das espécies foram submetidas ao processo de polpação kraft, considerando-se seis níveis de álcali ativo: 18,0; 19,5; 21,0; 22,5; 24,0 e 25,5%, mantendo-se constantes as outras variáveis de cozimento. Observou-se que o modelo de variação longitudinal para densidade básica da árvore mantém uma tendência dos valores serem mais elevados na base e valores mínimos entre 25 e 50% da altura comercial. Quanto à composição química os valores foram distintos para todas as espécies e apresentaram uma variação de 60,99% para A. auriculiformis (1) e 68,79% para A. aulacocarpa (1) como valores mínimo e máximo. Os resultados obtidos quanto às dimensões das fibras não apresentaram padrões definidos de variação entre as espécies. Notou-se quanto aos parâmetros da polpação um efeito diferenciado entre as espécies de Acacia para as distintas cargas de álcali ativo. Considerando a curva de polpação para obtenção de número kappa 18 entre as espécies estudadas, obteve-se como média para a demanda carga de alcalina 23,8% e a média do rendimento depurado nestas condições foi 54,72%. A espécie A. crassicarpa procedência (1) destacou-se por apresentar a maior conversão em massa (ton/ha) de madeira na produção de celulose e a A. mangium procedências (1, 2, 3 e principalmente 4) alcançaram os melhores resultados de rendimento frente ao processo kraft de polpação comparando toas as espécies deste estudo. / The genus Acacia includes fast-growing short-rotation (7 to 9 years) forest tree species of commercial interest to produce pulp, wood panels, and energy. In Brazil, the utilization of species of the genus Acacia to produce short fiber pulp is still highly restricted and few studies are available that demonstrate the potential use of this raw material to obtain pulp presenting the properties displayed by species of the genus Eucalyptus. This study aimed at analyzing the potential of some species of the genus Acacia (A. aulacocarpa, A. auriculiformis, A. crassicarpa, A. mangium) planted in Brazil to produce kraft pulp, considering the aspects related to wood and pulp quality, consumption of chemical reagents during the pulping process, yield, and cellulose pulp production capacity. Furthermore, this work searched for strategical information for the national pulp and paper sector as to possible threats and opportunities presented by materials not used in Brazil in order to provide knowledge and discuss the potential competitiveness of these types of wood in the world market for pulp and paper. To accomplish these goals, we used 11 5-year-old distinct genetic materials, each one of them considered a different treatment. The following traits were evaluated: basic density, longitudinal variation, fiber dimensions, chemical composition, and kraft pulping process parameters (total yield, screened yield, rejects content, kappa number and relations). To carry out these evaluations, each wood species underwent the kraft pulping process considering six levels of active alkalis: 18.0, 19.5, 21.0, 22.5, 24.0, and 25.5%, keeping all other cooking variables constant. We observed that the model of longitudinal variation in the basic density maintains the trend of reaching higher values in the base of the tree and minimum values between 25% and 50% of the commercial height. The values found for the chemical composition were different for all the species studied and the holocellulose content ranged from a minimum of 60.99% found for A. auriculiformis (1) to a maximum of 68.79% found for A. aulacocarpa (1). The results obtained for fiber dimensions did not present defined patterns of variation among the species. The kraft pulping process parameters showed differences among Acacia species for the distinct levels of active alkalis. Considering the pulping process curve to get the kappa number of 18 among the species studied, the average alkali demand was 23.8% and the average depurated yield under these conditions was 54.72%. A. crassicarpa procedure 1 presented the highest conversion into wood mass (t/ha) to produce cellulose and A. mangium procedures (1, 2, 3, and mainly 4) reached the best results in terms of yield when they underwent the kraft pulping process compared to all the species studied herein.
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Hybrid nanophotonic elements and sensing devices based on photonic crystal structuresBarth, Michael 12 July 2010 (has links)
Die vorliegende Forschungsarbeit widmet sich der Entwicklung und Untersuchung neuartiger photonischer Kristallstrukuren für Anwendungen in den Gebieten der Nanophotonik und Optofluidik. Dabei konzentriert sich eine erste Serie von Experimenten auf die Charakterisierung und Optimierung photonischer Kristallresonatoren im sichtbaren Spektralbereich, wobei bisher unerreichte Resonatorgüten von bis zu 3400 gezeigt werden können. Diese Strukturen werden anschließend als Plattformen zur Herstellung von hybriden nanophotonischen Bauelementen verwendet, indem externe Partikel (wie z.B. Diamant-Nanokristalle und Metall-Nanopartikel) in kontrollierter Art und Weise an die Resonatoren gekoppelt werden. Zu diesem Zweck wird eine Nanomanipulationsmethode entwickelt, welche Rastersonden zur gezielten Positionierung und Anordnung von Partikeln auf den photonischen Kristallstrukturen benutzt. Verschiedene Arten solcher Hybridelemente werden realisiert und untersucht, einschließlich diamant-gekoppelter Resonatoren, plasmon-gekoppelter Resonatoren und Metall-Diamant Hybridstrukturen. Außer für Anwendungen auf dem Gebiet der Nanophotonik werden verschiedene photonische Kristallstrukturen auch hinsichtlich ihres Leistungsvermögens als biochemische Sensorelemente erforscht. Zum ersten Mal wird eine umfassende numerische Analyse der optischen Kräfte auf Objekte im Nahfeld photonischer Kristallresonatoren durchgeführt, welche neue Möglichkeiten zum Einfang sowie zur Detektion und Untersuchung biologischer Partikel in integrierten optofluidischen Bauteilen bieten. Weiterhin werden unterschiedliche photonische Kristallfasern bezüglich ihrer Detektionssensitivität in Absorptions- und Fluoreszenzmessungen untersucht, wobei sich eine klare Überlegenheit von selektiv befüllten Hohlkern-Designs im Vergleich zu Festkern-Fasern offenbart. / This thesis deals with the development and investigation of novel photonic crystal structures for applications in nanophotonics and optofluidics. Thereby, a first series of experiments focuses on the characterization and optimization of photonic crystal cavities in the visible wavelength range, demonstrating unprecedented cavity quality factors of up to 3400. These structures are subsequently employed as platforms for the creation of advanced hybrid nanophotonic elements by coupling external particles (such as diamond nanocrystals and metal nanoparticles) to the cavities in a well-controlled manner. For this purpose, a nanomanipulation method is developed, utilizing scanning probes for the deterministic positioning and assembly of particles on the photonic crystal structures. Various types of such hybrid elements are realized and investigated, including diamond-coupled cavities, plasmon-coupled cavities, and metal-diamond hybrid structures. Apart from applications in nanophotonics, different types of photonic crystal structures are also studied with regard to their performance as biochemical sensing elements. For the first time a thorough numerical analysis of the optical forces exerted on objects in the near-field of photonic crystal cavities is conducted, providing novel means to trap, detect, and investigate biological particles in integrated optofluidic devices. Furthermore, various types of photonic crystal fibers are studied with regard to their detection sensitivity in absorption and fluorescence measurements, revealing a clear superiority of selectively infiltrated hollow-core designs in comparison to solid-core fibers.
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Qualidade da madeira de espécies do gênero Acacia plantadas no Brasil / Wood quality of species of the genus Acacia planted in BrazilGláucia Soares Barbosa de Alencar 09 June 2009 (has links)
O gênero Acacia apresenta espécies florestais de rápido crescimento, com rotações curtas (7 a 9 anos) e de grande interesse comercial para produção de polpa celulósica, painéis de madeira e energia. No Brasil a utilização de espécies do gênero Acacia na produção de celulose de fibra curta ainda é bastante restrita e são poucos os estudos que demonstram a potencialidade do uso dessa matéria-prima para a obtenção de polpa celulósica com as propriedades alcançadas pelas espécies do gênero Eucalyptus. Este trabalho teve como objetivo analisar o potencial de algumas espécies do gênero Acacia (A.aulacocarpa, A. auriculiformis, A. crassicarpa, A. mangium) plantados no Brasil, para a produção de polpa celulósica Kraft, considerando os aspectos relacionados à qualidade da madeira e da polpa, o consumo de reagentes químicos na polpação e também os aspectos relacionados ao rendimento e a capacidade de produção de polpa celulósica; adicionalmente buscou-se fornecer informações estratégicas para o setor celulósico nacional com relação as possíveis ameaças e as oportunidades apresentadas pelos materiais não utilizados no Brasil. Para realização deste trabalho foram utilizados 11 materiais genéticos distintos e cada um dos materiais genéticos foi considerado um tratamento e todos com 5 anos de idade. As características avaliadas foram: densidade básica, dimensões das fibras, composição química e parâmetros da polpação kraft (rendimento bruto, rendimento depurado, teor de rejeitos, número kappa e relações). Para este último objetivo, as madeiras de cada uma das espécies foram submetidas ao processo de polpação kraft, considerando-se seis níveis de álcali ativo: 18,0; 19,5; 21,0; 22,5; 24,0 e 25,5%, mantendo-se constantes as outras variáveis de cozimento. Observou-se que o modelo de variação longitudinal para densidade básica da árvore mantém uma tendência dos valores serem mais elevados na base e valores mínimos entre 25 e 50% da altura comercial. Quanto à composição química os valores foram distintos para todas as espécies e apresentaram uma variação de 60,99% para A. auriculiformis (1) e 68,79% para A. aulacocarpa (1) como valores mínimo e máximo. Os resultados obtidos quanto às dimensões das fibras não apresentaram padrões definidos de variação entre as espécies. Notou-se quanto aos parâmetros da polpação um efeito diferenciado entre as espécies de Acacia para as distintas cargas de álcali ativo. Considerando a curva de polpação para obtenção de número kappa 18 entre as espécies estudadas, obteve-se como média para a demanda carga de alcalina 23,8% e a média do rendimento depurado nestas condições foi 54,72%. A espécie A. crassicarpa procedência (1) destacou-se por apresentar a maior conversão em massa (ton/ha) de madeira na produção de celulose e a A. mangium procedências (1, 2, 3 e principalmente 4) alcançaram os melhores resultados de rendimento frente ao processo kraft de polpação comparando toas as espécies deste estudo. / The genus Acacia includes fast-growing short-rotation (7 to 9 years) forest tree species of commercial interest to produce pulp, wood panels, and energy. In Brazil, the utilization of species of the genus Acacia to produce short fiber pulp is still highly restricted and few studies are available that demonstrate the potential use of this raw material to obtain pulp presenting the properties displayed by species of the genus Eucalyptus. This study aimed at analyzing the potential of some species of the genus Acacia (A. aulacocarpa, A. auriculiformis, A. crassicarpa, A. mangium) planted in Brazil to produce kraft pulp, considering the aspects related to wood and pulp quality, consumption of chemical reagents during the pulping process, yield, and cellulose pulp production capacity. Furthermore, this work searched for strategical information for the national pulp and paper sector as to possible threats and opportunities presented by materials not used in Brazil in order to provide knowledge and discuss the potential competitiveness of these types of wood in the world market for pulp and paper. To accomplish these goals, we used 11 5-year-old distinct genetic materials, each one of them considered a different treatment. The following traits were evaluated: basic density, longitudinal variation, fiber dimensions, chemical composition, and kraft pulping process parameters (total yield, screened yield, rejects content, kappa number and relations). To carry out these evaluations, each wood species underwent the kraft pulping process considering six levels of active alkalis: 18.0, 19.5, 21.0, 22.5, 24.0, and 25.5%, keeping all other cooking variables constant. We observed that the model of longitudinal variation in the basic density maintains the trend of reaching higher values in the base of the tree and minimum values between 25% and 50% of the commercial height. The values found for the chemical composition were different for all the species studied and the holocellulose content ranged from a minimum of 60.99% found for A. auriculiformis (1) to a maximum of 68.79% found for A. aulacocarpa (1). The results obtained for fiber dimensions did not present defined patterns of variation among the species. The kraft pulping process parameters showed differences among Acacia species for the distinct levels of active alkalis. Considering the pulping process curve to get the kappa number of 18 among the species studied, the average alkali demand was 23.8% and the average depurated yield under these conditions was 54.72%. A. crassicarpa procedure 1 presented the highest conversion into wood mass (t/ha) to produce cellulose and A. mangium procedures (1, 2, 3, and mainly 4) reached the best results in terms of yield when they underwent the kraft pulping process compared to all the species studied herein.
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Improvement of the retention-fromation relationship using three-component retention aid systemsSvedberg, Anna January 2012 (has links)
QC 20120530
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Spegel, spegel på väggen där: : En studie kring balladens textliga funktion i kvinnliga traditionsbärares liv under 1800-taletHenriksson, Christina, Eriksson, Emma January 2006 (has links)
Emma Eriksson and Christina Henriksson: Mirror mirror on the wall: A study on how ballad texts reflect women as carriers of tradition in nineteenth century Sweden, Växjö: Musikvetenskap C, 10 p 2005. The aim of this essay is to analyze relations between ballad texts and the lives of female carriers of tradition in nineteenth century Sweden. Four carriers of tradition have been chosen, connected by being informants to Gunnar Olof Hyltén Cavallius, a collector of medieval ballads in Sweden around 1850. The four women are Anna Ehrenström (1786-1857), Catarina Andersdotter (1792-1857), Johanna Gustafva Angel (1791-1869) och Wendela Hebbe (1808-1899). Their lives are seen from a women’s historical perspective. Although there are uncertainties regarding to what extent the collected repertoire of the four women reflects their repertoire as a whole, some conclusions can be drawn regarding relations between remaining ballads and their lives as women. These relations are traced in textual themes and in textual form (e.g. repetition of words and phrases). The closeness between life situation, and choice and shape of ballad texts, leads us to conclude that these occurrences are due to deliberate choices, and that the ballad text thus mirror the lives of these women.
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Möglichkeiten und Grenzen bewegungstherapeutischer Intervention bei ParkinsonpatientenAugustijn, Julia 12 November 2012 (has links) (PDF)
Die posturale Instabilität zählt zu den am meisten beeinträchtigenden Symptomen der Parkinson-Krankheit. Die Störung der motorischen Gleichgewichtskontrolle ist progressiv im Verlauf und weder durch medikamentöse noch durch operative Methoden zufriedenstellend einzudämmen. In der Bewegungstherapie werden häufig Gleichgewichtsübungen empfohlen, um ein Fortschreiten der körperlichen Einschränkungen zu verringern. Der aktuelle wissenschaftliche Stand lässt allerdings eine Einschätzung zur Effektivität von Gleichgewichtstraining bei Parkinsonpatienten kaum zu. Dies ist u. a. auf einen Mangel an geeigneten Testverfahren zur Beurteilung der posturalen Instabilität zurückzuführen.
In der vorliegenden Untersuchung wurden die Auswirkungen eines 12-wöchigen Gleichgewichtstrainings bei Parkinsonpatienten auf die posturale Stabilität in einem umfassenden Testdesign, bestehend aus alltagsmotorischen, biomechanischen und subjektiven Testverfahren evaluiert.
In nahezu allen eingesetzten Testverfahren zeigte sich ein mehr oder weniger deutlicher Trend zu einer Verbesserung der posturalen Stabilität. Durch den Einsatz einer Testbatterie konnte somit insgesamt von einem positiven Einfluss eines Gleichgewichtstrainings auf die posturale Stabilität von Parkinsonpatienten ausgegangen werden.
Weiterhin werden zahlreiche positive Nebeneffekte bzgl. der allgemeinen Fitness, der psychischen und sozialen Situation durch ein zielgerichtetes Gruppentraining unter geschulter Anleitung vermutet.
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Investigation of biological macromolecules using atomic force microscope-based techniquesBippes, Christian Alexander 19 August 2009 (has links) (PDF)
The atomic force microscope (AFM) provides a powerful instrument for investigating and manipulating biological samples down to the subnanometer scale. In contrast to other microscopy methods, AFM does not require labeling, staining, nor fixation of samples and allows the specimen to be fully hydrated in buffer solution during the experiments. Moreover, AFM clearly compares in resolution to other techniques.
In general, the AFM can be operated in an imaging or a force spectroscopy mode. In the present work, advantage was taken of this versatility to investigate single biomolecules and biomolecular assemblies.
A novel approach to investigate the visco-elastic behavior of biomolecules under force was established, using dextran as an example. While a molecule tethered between a solid support and the cantilever tip was stretched at a constant velocity, the thermally driven oscillation of the cantilever was recorded. Analysis of the cantilever Brownian noise provided information about the visco-elastic properties of dextran that corresponded well to parameters obtained by alternative methods. However, the approach presented here was easier to implement and less time-consuming than previously used methods.
A computer controlled force-clamp system was set up, circumventing the need for custom built analogue electronics. A commercial PicoForce AFM was extended by two computers which hosted data acquisition hardware. While the first computer recorded data, the second computer drove the AFM bypassing the manufacturer's microscope control software. To do so, a software-based proportional-integral-differential (PID) controller was implemented on the second computer. It allowed the force applied to a molecule to be held constant over time. After tuning of the PID controller, response times obtained using that force-clamp setup were comparable to those of the recently reported analogue systems. The performance of the setup was demonstrated by force-clamp unfolding of a pentameric Ig25 construct and the membrane protein NhaA. In the latter case, short-lived unfolding intermediates that were populated for less than 10 ms, could be revealed.
Conventional single-molecule dynamic force spectroscopy was used to unfold the serine:threonine antiporter SteT from Bacillus subtilis, an integral membrane protein. Unfolding force patterns revealed the unfolding barriers stabilizing structural segments of SteT. Ligand binding did not induce new unfolding barriers suggesting that weak interactions with multiple structural segments were involved. In contrast, ligand binding caused changes in the energy landscape of all structural segments, thus turning the protein from a brittle, rigid into a more stable, structurally flexible conformation. Functionally, rigidity in the ligand-free state was thought to facilitate specific ligand binding, while flexibility and increased stability were required for conformational changes associated with substrate translocation. These results support the working model for transmembrane transport proteins that provide alternate access of the binding site to either face of the membrane.
Finally, high-resolution imaging was exploited to visualize the extracellular surface of Cx26 gap junction hemichannels (connexons). AFM topographs reveal pH-dependent structural changes of the extracellular connexon surface in presence of HEPES, an aminosulfonate compound. At low pH (< 6.5), connexons showed a narrow and shallow channel entrance, which represented the closed pore. Increasing pH values resulted in a gradual opening of the pore, which was reflected by increasing channel entrance widths and depths. At pH > 7.6 the pore was fully opened and the pore diameter and depth did not increase further. Importantly, coinciding with pore gating a slight rotation of the subunits was observed. In the absence of aminosulfonate compounds, such as HEPES, acidification did not affect pore diameters and depths, retaining the open state. Thus, the intracellular concentration of taurine, a naturally abundant aminosulfonate compound, might be used to tune gap junction sensitivity at low pH.
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