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Wang, W. (author)This thesis explores advanced computational techniques in super-resolution microscopy (SRM), with the primary goal of pushing the limits of achievable resolution towards the 1 nm scale. It includes developments in particle fusion algorithms, data analysis of complex biological structures, and exploration of the impact of molecular dipole...doctoral thesis 2024
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Tišma, M. (author)This thesis explores the mechanisms that underlie chromosome organization in bacteria. Bacteria are considered amongst the simplest living organisms on our planet. They lack the cellular organization found in other domains of life (Archaea or Eukaryotics) and often have simpler life cycles. Over the past decade, we gained increasing knowledge...doctoral thesis 2024
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Tišma, M. (author), Kaljević, Jovana (author), Gruber, Stephan (author), Le, Tung B.K. (author), Dekker, C. (author)Bacterial cells require DNA segregation machinery to properly distribute a genome to both daughter cells upon division. The most common system involved in chromosome and plasmid segregation in bacteria is the ParABS system. A core protein of this system - partition protein B (ParB) - regulates chromosome organization and chromosome...review 2024
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Klughammer, N. (author)In this thesis, we have introduced palladium zero-mode waveguides (Pd ZMWs) as a valuable tool for biophysics research and have showcased several applications thereof. We started by using them as single-molecule sensors for free translocations of fluorophores, proteins, and voltage-driven translocations of DNA. By studying salt-gradient driven...doctoral thesis 2023
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Ornago, L. (author)In this dissertation, we analyse the charge transport of nanoscale molecular junctions in mechanically controllable break junction (MCBJ) experiments. In particular, we focus on the characterization of molecular features going beyond the "single-peak" picture, that is, considering features in the measurements in addition to themost prominent one...doctoral thesis 2023
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Mc Cluskey, K.A. (author), Dekker, N.H. (author)TIRF (Total Internal Reflection Fluorescence) microscopy is a powerful tool for measuring the intra- and intermolecular dynamics of fluorescently-labeled single molecules. As TIRF measurements move to more complex biological systems with more fluorescent probes, the multi-band-pass dichroic that separates excitation from emission becomes...journal article 2023
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Yang, Sora (author), Klughammer, N. (author), Barth, A. (author), Tanenbaum, M. (author), Dekker, C. (author)Single-molecule fluorescence imaging experiments generally require sub-nanomolar protein concentrations to isolate single protein molecules, which makes such experiments challenging in live cells due to high intracellular protein concentrations. Here, we show that single-molecule observations can be achieved in live cells through a drastic...journal article 2023
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Fan, D. (author), Cnossen, J.P. (author), Hung, S. (author), Kromm, D. (author), Dekker, N.H. (author), Verbiest, G.J. (author), Smith, C.S. (author)To better understand the interactions between biological molecules, a high optical resolution in all three dimensions is crucial. The intrinsically lower axial resolution of microscopes however, is a limiting factor in fluorescence imaging, correspondingly in fluorescence based single molecule localization microscopy (SMLM). Here, we present...journal article 2023
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Kim, Eugene (author), Barth, R. (author), Dekker, C. (author)SMC (structural maintenance of chromosomes) protein complexes are an evolutionarily conserved family of motor proteins that hold sister chromatids together and fold genomes throughout the cell cycle by DNA loop extrusion. These complexes play a key role in a variety of functions in the packaging and regulation of chromosomes, and they have...review 2023
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Filius, M. (author), van Wee, R.G. (author), Joo, C. (author)Fluorescence resonance energy transfer (FRET) is a photophysical phenomenon that has been repurposed as a biophysical tool to measure nanometer distances. With FRET by DNA eXchange, or FRET X, many points of interest (POIs) in a single object can be probed, overcoming a major limitation of conventional single-molecule FRET. In FRET X, short...book chapter 2023
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Ploetz, Evelyn (author), Ambrose, Benjamin (author), Barth, A. (author), Börner, Richard (author), Kapanidis, Achillefs N. (author), Lohman, Timothy M. (author), Steffen, Fabio D. (author), Cordes, Thorben (author), Magennis, Steven W. (author), Lerner, Eitan (author)PIFE was first used as an acronym for protein-induced fluorescence enhancement, which refers to the increase in fluorescence observed upon the interaction of a fluorophore, such as a cyanine, with a protein. This fluorescence enhancement is due to changes in the rate ofcis/transphotoisomerisation. It is clear now that this mechanism is...review 2023
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Hsu, C. (author)This dissertation presents the experimental work on singlemolecule charge transport, particularly on the aspects of spin and orbital conformational effects in singlemolecule junctions. In this dissertation, we employ and develop various singlemolecule techniques, the mechanically controlled break junction (MCBJ), thermoelectric electromigrated...doctoral thesis 2022
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Filius, M. (author)Proteins are the workhorses of the cell, as such, they form the basis of all living systems. In order to fully understand biological processes, the ability to identify and quantify the proteins in cells is crucial. Identification can be achieved by determining the amino acid sequence of proteins, since this sequence is unique for each protein....doctoral thesis 2022
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Wen, C. (author), Bertosin, E. (author), Shi, X. (author), Dekker, C. (author), Schmid, Sonja (author)Nanopores are versatile single-molecule sensors offering a simple label-free readout with great sensitivity. We recently introduced the nanopore electro-osmotic trap (NEOtrap) which can trap and sense single unmodified proteins for long times. The trapping is achieved by the electro-osmotic flow (EOF) generated from a DNA-origami sphere...journal article 2022
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Janissen, R. (author), Eslami Mossallam, B. (author), Artsimovitch, Irina (author), Depken, S.M. (author), Dekker, N.H. (author)Pausing by bacterial RNA polymerase (RNAp) is vital in the recruitment of regulatory factors, RNA folding, and coupled translation. While backtracking and intra-structural isomerization have been proposed to trigger pausing, our mechanistic understanding of backtrack-associated pauses and catalytic recovery remains incomplete. Using high...journal article 2022
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Severins, I.W.H. (author), Joo, C. (author), van Noort, John (author)Next-generation sequencing techniques have led to a new quantitative dimension in the biological sciences. In particular, integrating sequencing techniques with biophysical tools allows sequence-dependent mechanistic studies. Using the millions of DNA clusters that are generated during sequencing to perform high-throughput binding affinity...review 2022
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Kuijpers, L.C. (author), van Laar, T. (author), Janissen, R. (author), Dekker, N.H. (author)Multiplexed single-molecule magnetic tweezers (MT) have recently been employed to probe the RNA synthesis dynamics of RNA-dependent RNA polymerases (RdRp). Here, we present a protocol for simultaneously probing the RNA synthesis dynamics of hundreds of single polymerases with MT. We describe the preparation of a dsRNA construct for probing...journal article 2022
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Hulleman, C.N. (author)<i>Localization microscopy has circumvented the diffraction limit by sequentially</i> imaging individual light emitting molecules at a time. The position of these individual molecules can be determined and a super-resolution reconstruction is made with improved resolution. Normally freely rotating emitters are used such that the point spread...doctoral thesis 2021
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Bera, Subhas Chandra (author), Seifert, Mona (author), Kirchdoerfer, Robert N. (author), van Nies, Pauline (author), Wubulikasimu, Yibulayin (author), Quack, Salina (author), Papini, Flávia S. (author), Depken, S.M. (author), Dulin, D. (author)Coronaviruses have evolved elaborate multisubunit machines to replicate and transcribe their genomes. Central to these machines are the RNA-dependent RNA polymerase subunit (nsp12) and its intimately associated cofactors (nsp7 and nsp8). We use a high-throughput magnetic-tweezers approach to develop a mechanochemical description of this core...journal article 2021
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Navikas, Vytautas (author), Descloux, Adrien C. (author), Grußmayer, K.S. (author), Marion, Sanjin (author), Radenovic, Aleksandra (author)A variety of modern super-resolution microscopy methods provide researchers with previously inconceivable biological sample imaging opportunities at a molecular resolution. All of these techniques excel at imaging samples that are close to the coverslip, however imaging at large depths remains a challenge due to aberrations caused by the...journal article 2021