Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Quantifying absolute addressability in DNA origami with molecular resolution.

ABSTRACT: Self-assembled DNA nanostructures feature an unprecedented addressability with sub-nanometer precision and accuracy. This addressability relies on the ability to attach functional entities to single DNA strands in these structures. The efficiency of this attachment depends on two factors: incorporation of the strand of interest and accessibility of this strand for downstream modification. Here we use DNA-PAINT super-resolution microscopy to quantify both incorporation and accessibility of all individual strands in DNA origami with molecular resolution. We find that strand incorporation strongly correlates with the position in the structure, ranging from a minimum of 48% on the edges to a maximum of 95% in the center. Our method offers a direct feedback for the rational refinement of the design and assembly process of DNA nanostructures and provides a long sought-after quantitative explanation for efficiencies of DNA-based nanomachines.

SUBMITTER: Strauss MT

PROVIDER: S-EPMC5913233 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Quantifying absolute addressability in DNA origami with molecular resolution.

Strauss Maximilian T MT Schueder Florian F Haas Daniel D Nickels Philipp C PC Jungmann Ralf R

Nature communications 20180423 1

Self-assembled DNA nanostructures feature an unprecedented addressability with sub-nanometer precision and accuracy. This addressability relies on the ability to attach functional entities to single DNA strands in these structures. The efficiency of this attachment depends on two factors: incorporation of the strand of interest and accessibility of this strand for downstream modification. Here we use DNA-PAINT super-resolution microscopy to quantify both incorporation and accessibility of all in ...[more]

PMID: 29686288

Similar Datasets

A DNA origami platform for quantifying protein copy number in super-resolution.

Project description:Single-molecule-based super-resolution microscopy offers researchers a unique opportunity to quantify protein copy number with nanoscale resolution. However, while fluorescent proteins have been characterized for quantitative imaging using calibration standards, similar calibration tools for immunofluorescence with small organic fluorophores are lacking. Here we show that DNA origami, in combination with GFP antibodies, is a versatile platform for calibrating fluorophore and antibody labeling efficiency to quantify protein copy number in cellular contexts using super-resolution microscopy.

| S-EPMC5534338 | biostudies-literature

Photocontrolled Dopamine Polymerization on DNA Origami with Nanometer Resolution.

Project description:Temporal and spatial control over polydopamine formation on the nanoscale can be achieved by installing an irradiation-sensitive polymerization system on DNA origami. Precisely distributed G-quadruplex structures on the DNA template serve as anchors for embedding the photosensitizer protoporphyrin IX, which-upon irradiation with visible light-induces the multistep oxidation of dopamine to polydopamine, producing polymeric structures on designated areas within the origami framework. The photochemical polymerization process allows exclusive control over polydopamine layer formation through the simple on/off switching of the light source. The obtained polymer-DNA hybrid material shows significantly enhanced stability, paving the way for biomedical and chemical applications that are typically not possible owing to the sensitivity of DNA.

| S-EPMC7186833 | biostudies-literature

Quantification of Strand Accessibility in Biostable DNA Origami with Single-Staple Resolution.

Project description:DNA-based nanostructures are actively gaining interest as tools for biomedical and therapeutic applications following the recent development of protective coating strategies prolonging structural integrity in physiological conditions. For tailored biological action, these nanostructures are often functionalized with targeting or imaging labels using DNA base pairing. Only if these labels are accessible on the structure's surface will they be able to interact with their intended biological target. However, the accessibility of functional sites for different geometries and environments, specifically after the application of a protective coating, is currently not known. Here, we assay this accessibility on the level of single handle strands with two- and three-dimensional resolution using DNA-PAINT and show that the hybridization kinetics of top and bottom sides on the same nanostructure linked to a surface remain unaltered. We furthermore demonstrate that the functionality of the structures remains available after an oligolysine-PEG coating is applied, enabling bioassays where functionality and stability are imperative.

| S-EPMC8613912 | biostudies-literature

DNA Origami Reorganizes upon Interaction with Graphite: Implications for High-Resolution DNA Directed Protein Patterning.

Project description:Although there is a long history of the study of the interaction of DNA with carbon surfaces, limited information exists regarding the interaction of complex DNA-based nanostructures with the important material graphite, which is closely related to graphene. In view of the capacity of DNA to direct the assembly of proteins and optical and electronic nanoparticles, the potential for combining DNA-based materials with graphite, which is an ultra-flat, conductive carbon substrate, requires evaluation. A series of imaging studies utilizing Atomic Force Microscopy has been applied in order to provide a unified picture of this important interaction of structured DNA and graphite. For the test structure examined, we observe a rapid destabilization of the complex DNA origami structure, consistent with a strong interaction of single-stranded DNA with the carbon surface. This destabilizing interaction can be obscured by an intentional or unintentional primary intervening layer of single-stranded DNA. Because the interaction of origami with graphite is not completely dissociative, and because the frustrated, expanded structure is relatively stable over time in solution, it is demonstrated that organized structures of pairs of the model protein streptavidin can be produced on carbon surfaces using DNA origami as the directing material.

| S-EPMC5245763 | biostudies-literature

Toward Absolute Molecular Numbers in DNA-PAINT.

Project description:Single-molecule localization microscopy (SMLM) has revolutionized optical microscopy, extending resolution down to the level of individual molecules. However, the actual counting of molecules relies on preliminary knowledge of the blinking behavior of individual targets or on a calibration to a reference. In particular for biological applications, great care has to be taken because a plethora of factors influence the quality and applicability of calibration-dependent approaches to count targets in localization clusters particularly in SMLM data obtained from heterogeneous samples. Here, we present localization-based fluorescence correlation spectroscopy (lbFCS) as the first absolute molecular counting approach for DNA-points accumulation for imaging in nanoscale topography (PAINT) microscopy and, to our knowledge, for SMLM in general. We demonstrate that lbFCS overcomes the limitation of previous DNA-PAINT counting and allows the quantification of target molecules independent of the localization cluster density. In accordance with the promising results of our systematic proof-of-principle study on DNA origami structures as idealized targets, lbFCS could potentially also provide quantitative access to more challenging biological targets featuring heterogeneous cluster sizes in the future.

| S-EPMC6856960 | biostudies-literature

Molecular behavior of DNA origami in higher-order self-assembly.

Project description:DNA-based self-assembly is a unique method for achieving higher-order molecular architectures made possible by the fact that DNA is a programmable information-coding polymer. In the past decade, two main types of DNA nanostructures have been developed: branch-shaped DNA tiles with small dimensions (commonly up to ∼20 nm) and DNA origami tiles with larger dimensions (up to ∼100 nm). Here we aimed to determine the important factors involved in the assembly of DNA origami superstructures. We constructed a new series of rectangular-shaped DNA origami tiles in which parallel DNA helices are arranged in a zigzag pattern when viewed along the DNA helical axis, a design conceived in order to relax an intrinsic global twist found in the original planar, rectangular origami tiles. Self-associating zigzag tiles were found to form linear arrays in both diagonal directions, while planar tiles showed significant growth in only one direction. Although the series of zigzag tiles were designed to promote two-dimensional array formation, one-dimensional linear arrays and tubular structures were observed instead. We discovered that the dimensional aspect ratio of the origami unit tiles and intertile connection design play important roles in determining the final products, as revealed by atomic force microscopy imaging. This study provides insight into the formation of higher-order structures from self-assembling DNA origami tiles, revealing their unique behavior in comparison with conventional DNA tiles having smaller dimensions.

| S-EPMC3071357 | biostudies-literature

Quantifying molecular bias in DNA data storage.

Project description:DNA has recently emerged as an attractive medium for archival data storage. Recent work has demonstrated proof-of-principle prototype systems; however, very uneven (biased) sequencing coverage has been reported, which indicates inefficiencies in the storage process. Deviations from the average coverage in the sequence copy distribution can either cause wasteful provisioning in sequencing or excessive number of missing sequences. Here, we use millions of unique sequences from a DNA-based digital data archival system to study the oligonucleotide copy unevenness problem and show that the two paramount sources of bias are the synthesis and amplification (PCR) processes. Based on these findings, we develop a statistical model for each molecular process as well as the overall process. We further use our model to explore the trade-offs between synthesis bias, storage physical density, logical redundancy, and sequencing redundancy, providing insights for engineering efficient, robust DNA data storage systems.

| S-EPMC7324401 | biostudies-literature

DNA origami protection and molecular interfacing through engineered sequence-defined peptoids.

Project description:DNA nanotechnology has established approaches for designing programmable and precisely controlled nanoscale architectures through specific Watson-Crick base-pairing, molecular plasticity, and intermolecular connectivity. In particular, superior control over DNA origami structures could be beneficial for biomedical applications, including biosensing, in vivo imaging, and drug and gene delivery. However, protecting DNA origami structures in complex biological fluids while preserving their structural characteristics remains a major challenge for enabling these applications. Here, we developed a class of structurally well-defined peptoids to protect DNA origamis in ionic and bioactive conditions and systematically explored the effects of peptoid architecture and sequence dependency on DNA origami stability. The applicability of this approach for drug delivery, bioimaging, and cell targeting was also demonstrated. A series of peptoids (PE1-9) with two types of architectures, termed as "brush" and "block," were built from positively charged monomers and neutral oligo-ethyleneoxy monomers, where certain designs were found to greatly enhance the stability of DNA origami. Through experimental and molecular dynamics studies, we demonstrated the role of sequence-dependent electrostatic interactions of peptoids with the DNA backbone. We showed that octahedral DNA origamis coated with peptoid (PE2) can be used as carriers for anticancer drug and protein, where the peptoid modulated the rate of drug release and prolonged protein stability against proteolytic hydrolysis. Finally, we synthesized two alkyne-modified peptoids (PE8 and PE9), conjugated with fluorophore and antibody, to make stable DNA origamis with imaging and cell-targeting capabilities. Our results demonstrate an approach toward functional and physiologically stable DNA origami for biomedical applications.

| S-EPMC7104344 | biostudies-literature

Molecular force spectroscopy with a DNA origami-based nanoscopic force clamp.

Project description:Forces in biological systems are typically investigated at the single-molecule level with atomic force microscopy or optical and magnetic tweezers, but these techniques suffer from limited data throughput and their requirement for a physical connection to the macroscopic world. We introduce a self-assembled nanoscopic force clamp built from DNA that operates autonomously and allows massive parallelization. Single-stranded DNA sections of an origami structure acted as entropic springs and exerted controlled tension in the low piconewton range on a molecular system, whose conformational transitions were monitored by single-molecule Förster resonance energy transfer. We used the conformer switching of a Holliday junction as a benchmark and studied the TATA-binding protein-induced bending of a DNA duplex under tension. The observed suppression of bending above 10 piconewtons provides further evidence of mechanosensitivity in gene regulation.

| S-EPMC6546592 | biostudies-literature

Super-Resolution Imaging Conditions for enhanced Yellow Fluorescent Protein (eYFP) Demonstrated on DNA Origami Nanorulers.

Project description:Photostability is one of the crucial properties of a fluorophore which strongly influences the quality of single molecule-based super-resolution imaging. Enhanced yellow fluorescent protein (eYFP) is one of the most widely used versions of fluorescent proteins in modern cell biology exhibiting fast intrinsic blinking and reversible photoactivation by UV light. Here, we developed an assay for studying photostabilization of single eYFP molecules with respect to fast blinking and demonstrated a 6-fold enhanced photostability of single eYFP molecules with a beneficial influence on the blinking kinetics under oxygen removal and addition of aliphatic thiols (dSTORM-buffer). Conjugation to single stranded DNA and immobilization via DNA hybridization on a DNA origami 12 helix bundle in aqueous solution allowed photophyiscal studies of eYFP at the single-molecule level and at close to physiological conditions. The benefit of improved photophysical properties for localization-based super-resolution microscopy is demonstrated and quantitatively characterized by imaging 12 helix bundle DNA origami nanorulers with binding sites at designed distances of 160 and 100 nm and by imaging microtubules in fixed mammalian Vero cells.

| S-EPMC4571581 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data