@phdthesis{Knigge2020,
  author    = {Knigge, Xenia},
  title     = {Einzelmolek{\"u}l-Manipulation mittels Nano-Elektroden und Dielektrophorese},
  doi       = {10.25932/publishup-44313},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-443137},
  school      = {Universit{\"a}t Potsdam},
  pages     = {106, xxxii},
  year      = {2020},
  abstract  = {In dieser Arbeit wurden Nano-Elektroden-Arrays zur Einzel-Objekt-Immobilisierung mittels Dielektrophorese verwendet. Hierbei wurden fluoreszenzmarkierte Nano-Sph{\"a}ren als Modellsystem untersucht und die gewonnenen Ergebnisse auf biologische Proben {\"u}bertragen. Die Untersuchungen in Kombination mit verschiedenen Elektrodenlayouts f{\"u}hrten zu einer deterministischen Vereinzelung der Nano-Sph{\"a}ren ab einem festen Gr{\"o}ßenverh{\"a}ltnis zwischen Nano-Sph{\"a}re und Durchmesser der Elektrodenspitzen. An den Proteinen BSA und R-PE konnte eine dielektrophoretische Immobilisierung ebenfalls demonstriert und R-PE Molek{\"u}le zur Vereinzelung gebracht werden. Hierf{\"u}r war neben einem optimierten Elektrodenlayout, das durch Feldsimulationen den Feldgradienten betreffend gesucht wurde, eine Optimierung der Feldparameter, insbesondere von Spannung und Frequenz, erforderlich. Neben der Dielektrophorese erfolgten auch Beobachtungen anderer Effekte des elektrischen Feldes, wie z.B. Elektrolyse an Nano-Elektroden und Str{\"o}mungen {\"u}ber dem Elektroden-Array, hervorgerufen durch Joulesche W{\"a}rme und AC-elektroosmotischen Fluss. Zudem konnte Dielektrophorese an Silberpartikeln beobachtet werden und mittels Fluoreszenz-, Atom-Kraft-, Raster-Elektronen-Mikroskopie und energiedispersiver R{\"o}ntgenspektroskopie untersucht werden. Schließlich wurden die verwendeten Objektive und Kameras auf ihre Lichtempfindlichkeit hin analysiert, so dass die Vereinzelung von Biomolek{\"u}len an Nano-Elektroden nachweisbar war. Festzuhalten bleibt also, dass die Vereinzelung von Nano-Objekten und Biomolek{\"u}len an Nano-Elektroden-Arrays gelungen ist. Durch den parallelen Ansatz erlaubt dies, Aussagen {\"u}ber das Verhalten von Einzelmolek{\"u}len mit guter Statistik zu treffen.},
  language  = {de}
}
@phdthesis{Niedermayer2012,
  author    = {Niedermayer, Thomas},
  title     = {On the depolymerization of actin filaments},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus-63605},
  school      = {Universit{\"a}t Potsdam},
  year      = {2012},
  abstract  = {Actin is one of the most abundant and highly conserved proteins in eukaryotic cells. The globular protein assembles into long filaments, which form a variety of different networks within the cytoskeleton. The dynamic reorganization of these networks - which is pivotal for cell motility, cell adhesion, and cell division - is based on cycles of polymerization (assembly) and depolymerization (disassembly) of actin filaments. Actin binds ATP and within the filament, actin-bound ATP is hydrolyzed into ADP on a time scale of a few minutes. As ADP-actin dissociates faster from the filament ends than ATP-actin, the filament becomes less stable as it grows older. Recent single filament experiments, where abrupt dynamical changes during filament depolymerization have been observed, suggest the opposite behavior, however, namely that the actin filaments become increasingly stable with time. Several mechanisms for this stabilization have been proposed, ranging from structural transitions of the whole filament to surface attachment of the filament ends. The key issue of this thesis is to elucidate the unexpected interruptions of depolymerization by a combination of experimental and theoretical studies. In new depolymerization experiments on single filaments, we confirm that filaments cease to shrink in an abrupt manner and determine the time from the initiation of depolymerization until the occurrence of the first interruption. This duration differs from filament to filament and represents a stochastic variable. We consider various hypothetical mechanisms that may cause the observed interruptions. These mechanisms cannot be distinguished directly, but they give rise to distinct distributions of the time until the first interruption, which we compute by modeling the underlying stochastic processes. A comparison with the measured distribution reveals that the sudden truncation of the shrinkage process neither arises from blocking of the ends nor from a collective transition of the whole filament. Instead, we predict a local transition process occurring at random sites within the filament. The combination of additional experimental findings and our theoretical approach confirms the notion of a local transition mechanism and identifies the transition as the photo-induced formation of an actin dimer within the filaments. Unlabeled actin filaments do not exhibit pauses, which implies that, in vivo, older filaments become destabilized by ATP hydrolysis. This destabilization can be identified with an acceleration of the depolymerization prior to the interruption. In the final part of this thesis, we theoretically analyze this acceleration to infer the mechanism of ATP hydrolysis. We show that the rate of ATP hydrolysis is constant within the filament, corresponding to a random as opposed to a vectorial hydrolysis mechanism.},
  language  = {en}
}