Single-Particle Characterization of Aβ Oligomers in Solution

Yusko, Erik C.; Prangkio, Panchika; Sept, David; Rollings, Ryan; Li, Jiali; Mayer, Michael

doi:10.1021/nn300542q

Cited by 107 publications

(129 citation statements)

References 95 publications

Supporting

Mentioning

124

Contrasting

Unclassified

Order By: Relevance

“…17,21–24 One such material is DNA origami— an object obtained by folding a long strand of DNA into a predefined pattern. 25 Since the DNA origami technique was first demonstrated in 2006, it has been used to assemble a variety of complex three-dimensional objects.…”

mentioning

confidence: 99%

Ionic Conductivity, Structural Deformation, and Programmable Anisotropy of DNA Origami in Electric Field

et al. 2015

View full text Add to dashboard Cite

The DNA origami technique can enable functionalization of inorganic structures for single-molecule electric current recordings. Experiments have shown that several layers of DNA molecules—a DNA origami plate— placed on top of a solid-state nanopore is permeable to ions. Here, we report a comprehensive characterization of the ionic conductivity of DNA origami plates by means of all-atom molecular dynamics (MD) simulations and nanocapillary electric current recordings. Using the MD method, we characterize the ionic conductivity of several origami constructs, revealing the local distribution of ions, the distribution of the electrostatic potential and contribution of different molecular species to the current. The simulations determine the dependence of the ionic conductivity on the applied voltage, the number of DNA layers, the nucleotide content and the lattice type of the plates. We demonstrate that increasing the concentration of Mg2+ ions makes the origami plates more compact, reducing their conductivity. The conductance of a DNA origami plate on top of a solid-state nanopore is determined by the two competing effects: bending of the DNA origami plate that reduces the current and separation of the DNA origami layers that increases the current. The latter is produced by the electro-osmotic flow and is reversible at the time scale of a hundred nanoseconds. The conductance of a DNA origami object is found to depend on its orientation, reaching maximum when the electric field aligns with the direction of the DNA helices. Our work demonstrates feasibility of programming the electrical properties of a self-assembled nanoscale object using DNA.

show abstract

mentioning

confidence: 99%

Ionic Conductivity, Structural Deformation, and Programmable Anisotropy of DNA Origami in Electric Field

et al. 2015

View full text Add to dashboard Cite

show abstract

“…17–19 As a single molecule analytical tool, nanopores have been used to investigate the biochemical and biophysical properties of proteins, e.g. folding 20 and unfolding, 21–23 protein aggregation, 24 ubiquitin linkage type, 25 and enzymatic activity 17, 26 . Recently, the unfolding and threading of a protein through a nanopore by an unfoldase demonstrated the unidirectional and processive translocation of a linearized peptide strand which opens the possibility for nanopore-based protein sequencing.…”

mentioning

confidence: 99%

Electrostatic Interactions between OmpG Nanopore and Analyte Protein Surface Can Distinguish between Glycosylated Isoforms

Fahie

Chen

2015

J. Phys. Chem. B

View full text Add to dashboard Cite

The flexible loops decorating the entrance of OmpG nanopore move dynamically during ionic current recording. The gating caused by these flexible loops changes when a target protein is bound. The gating is characterized by parameters including frequency, duration and open-pore current and these features combine to reveal the identity of a specific analyte protein. Here, we show that OmpG nanopore equipped with a biotin ligand can distinguish glycosylated and deglycosylated isoforms of avidin by their differences in surface charge. Our studies demonstrate that the direct interaction between the nanopore and analyte surface, induced by the electrostatic attraction between the two molecules, are essential for protein isoform detection. Our technique is remarkably sensitive to the analyte surface, which may provide a useful tool for glycoprotein profiling.

show abstract

“…From these changes in the ionic current, one can extract information of the analyte molecular properties such as size, charge, conformational states and interactions with other biomolecules 22 . Both biological and solid-state nanopores have been used to study protein folding at the single-molecule level, revealing the conformational change and dynamics during protein unfolding [23][24][25][26] , and have also been used to observe macromolecular changes of proteins 27 . Quartz nanopipettes, a sub-class of solid-state nanopores, are low-cost and straightforward to fabricate, circumventing the technical barrier of using conventional and expensive solidstate nanopores or biological nanopores.…”

Section: Introductionmentioning

confidence: 99%

Single-molecule nanopore sensing of actin dynamics and drug binding

Wang

Wilkinson

Lin

et al. 2019

Preprint

View full text Add to dashboard Cite

Actin is a key protein in the dynamic processes within the eukaryotic cell. To date, methods exploring the molecular state of actin are limited to insights gained from structural approaches, providing a snapshot of protein folding, or methods that require chemical modifications compromising actin monomer thermostability. Nanopore sensing permits label-free investigation of native proteins and is ideally suited to study proteins such as actin that require specialised buffers and cofactors. Using nanopores we determined the state of actin at the macromolecular level (filamentous or globular) and in its monomeric form bound to inhibitors. We revealed urea-dependent and voltage-dependent transitional states and observed unfolding process within which sub-populations of transient actin oligomers are visible. We detected, in real-time, drug-binding and filament-growth events at the single-molecule level. This enabled us to calculate binding stoichiometries and to propose a model for protein dynamics using unmodified, native actin molecules, demostrating the promise of nanopores sensing for in-depth understanding of protein folding landscapes and for drug discovery.through different modes of action. Finally, we demonstrate the ability of nanopipette-based nanopores in drug discovery by measuring real-time drug-binding at a single molecule level, calculating the observed rate constant and the saturation concentration of Swinholide A dimer formation. Using these measurements, we propose a positively cooperative actin-binding model for the Swinholide A drug's mode of action. The ability to perform such studies, label-free and with single-molecule resolution demonstrates the potential of using quartz nanopipettes in both the direct probing of the unfolding of complex biomolecules but also for future molecular diagnostics and drug discovery.6 Results Experimental configurationNanopipettes were fabricated using laser-assisted pulling as reported previously 28 . The pulled nanopipettes terminated with a single nanopore with an average diameter of 25 ± 4 nm (N = 5), as measured by Scanning Electron Microscopy (Fig. 1a and Fig. S1). Linear IV curves were obtained in the monomeric actin buffer, and the nanopore conductance was determined to be 37.0 ± 3.9 nS (Fig. S2). Given the high potassium chloride concentration (1 M), we confirmed that actin remained monomeric in the nanopore buffer (Fig. S3). In all experiments, the analyte was introduced into an external reservoir (CIS) along with a reference/ground Ag/AgCl electrode. The pipette was filled only with buffer solution, and a patch/working Ag/AgCl electrode was inserted ( Fig. 1a). When an external electric field is applied, protein transport through the nanopore is a result of cooperative and competitive contributions from diffusion, electrophoretic (EP) and electroosmotic (EO) flows 29 . At pH 8.0, both the surface of the nanopipettes and actin (isoelectric point ~5.3) are negatively charged, meaning we see no translocation events at a negative voltage (Fig. S4). We used rel...

show abstract

Single-Particle Characterization of Aβ Oligomers in Solution

Cited by 107 publications

References 95 publications

Ionic Conductivity, Structural Deformation, and Programmable Anisotropy of DNA Origami in Electric Field

Ionic Conductivity, Structural Deformation, and Programmable Anisotropy of DNA Origami in Electric Field

Electrostatic Interactions between OmpG Nanopore and Analyte Protein Surface Can Distinguish between Glycosylated Isoforms

Single-molecule nanopore sensing of actin dynamics and drug binding

Contact Info

Product

Resources

About