Monitoring patterned enzymatic polymerization on DNA origami at single-molecule level

Nanoscale ◽  
2015 ◽  
Vol 7 (25) ◽  
pp. 10970-10973 ◽  
Author(s):  
A. H. Okholm ◽  
H. Aslan ◽  
F. Besenbacher ◽  
M. Dong ◽  
J. Kjems
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Dna Origami ◽  
Enzymatic Polymerization ◽  
Dna Polymerization ◽  
Single Molecule Level ◽  
Site Selective

Site-selective enzymatic polymerization by TdT on surface-bound DNA origami embedded in BSA. DNA polymerization was visualized real time by AFM.

DNA Origami as a DNA Repair Nanosensor at the Single-Molecule Level

2013 ◽  
Vol 52 (30) ◽  
pp. 7747-7750 ◽  
Author(s):  
Maria Tintoré ◽  
Isaac Gállego ◽  
Brendan Manning ◽  
Ramon Eritja ◽  
Carme Fàbrega
Keyword(s):  
Dna Repair ◽  
Single Molecule ◽  
Dna Origami ◽  
Single Molecule Level

scholarly journals Real-Time 3D Imaging at the Single Molecule Level of Signal Transduction in Saccharomyces CerevisiÆ Responding to Environmental Changes

2016 ◽  
Vol 110 (3) ◽  
pp. 145a
Author(s):  
Erik G. Hedlund ◽  
Sviatlana Shashkova ◽  
Adam J.M. Wollman ◽  
Stefan Hohmann ◽  
Mark C. Leake
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Signal Transduction ◽  
Real Time ◽  
Single Molecule ◽  
3D Imaging ◽  
Environmental Changes ◽  
Single Molecule Level

DNA Origami as a DNA Repair Nanosensor at the Single-Molecule Level

2013 ◽  
Vol 125 (30) ◽  
pp. 7901-7904 ◽  
Author(s):  
Maria Tintoré ◽  
Isaac Gállego ◽  
Brendan Manning ◽  
Ramon Eritja ◽  
Carme Fàbrega
Keyword(s):  
Dna Repair ◽  
Single Molecule ◽  
Dna Origami ◽  
Single Molecule Level

scholarly journals FluoSim: simulator of single molecule dynamics for fluorescence live-cell and super-resolution imaging of membrane proteins

2020 ◽  
Author(s):  
Matthieu Lagardère ◽  
Ingrid Chamma ◽  
Emmanuel Bouilhol ◽  
Macha Nikolski ◽  
Olivier Thoumine
Keyword(s):  
Real Time ◽  
Protein Dynamics ◽  
Single Molecule ◽  
Imaging Techniques ◽  
Simulated Data ◽  
Super Resolution ◽  
Molecular Complexes ◽  
Live Cell ◽  
Single Molecule Level ◽  
Resolution Imaging

AbstractFluorescence live-cell and super-resolution microscopy methods have considerably advanced our understanding of the dynamics and mesoscale organization of macro-molecular complexes that drive cellular functions. However, different imaging techniques can provide quite disparate information about protein motion and organization, owing to their respective experimental ranges and limitations. To address these limitations, we present here a unified computer program that allows one to model and predict membrane protein dynamics at the ensemble and single molecule level, so as to reconcile imaging paradigms and quantitatively characterize protein behavior in complex cellular environments. FluoSim is an interactive real-time simulator of protein dynamics for live-cell imaging methods including SPT, FRAP, PAF, and FCS, and super-resolution imaging techniques such as PALM, dSTORM, and uPAINT. The software, thoroughly validated against experimental data on the canonical neurexin-neuroligin adhesion complex, integrates diffusion coefficients, binding rates, and fluorophore photo-physics to calculate in real time the distribution of thousands of independent molecules in 2D cellular geometries, providing simulated data of protein dynamics and localization directly comparable to actual experiments.

scholarly journals Local, Single-Molecule Oxidative Cleaving of DNA Origami by C60 on an AFM Tip

2021 ◽  
Author(s):  
Ankita Ray ◽  
Cristiana Passiu ◽  
S.N Ramakrishna ◽  
Antonella Rossi ◽  
Akinori Kuzuya ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Damage ◽  
Single Molecule ◽  
Dna Cleavage ◽  
Dna Origami ◽  
Oxygen Species ◽  
Mica Surface ◽  
Single Molecule Level ◽  
Afm Tips ◽  
Morphology Changes

Spatially controlled single-molecule oxidation of DNA was performed by photocatalytic generation of singlet oxygen on chemically functionalized AFM tips. A waffle-type DNA origami deposited on a mica surface is site-specifically destroyed by generation of reactive oxygen species at the AFM tip, which was modified with C<sub>60</sub>-tripod photocatalyst. Upon AFM scanning under photoirradiation, DNA morphology changes, corresponding to oxidative damage were clearly observed at the single-molecule level. The DNA cleavage occurred with strict dependence on photoirradiation and the presence of C<sub>60 </sub>on the AFM tip.

DNA origami-based single-molecule force spectroscopy unravels the molecular basis of RNA Polymerase III pre-initiation complex stability

2019 ◽  
Author(s):  
Kevin Kramm ◽  
Tim Schröder ◽  
Jerome Gouge ◽  
Andrés Manuel Vera ◽  
Florian B. Heiss ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Single Molecule ◽  
Rna Polymerase Iii ◽  
Dna Origami ◽  
Initiation Complex ◽  
Single Molecule Level ◽  
Initiation Factors ◽  
Strict Requirement ◽  
Rnap Ii ◽  
Pol Iii

AbstractThe TATA-binding protein (TBP) and a transcription factor (TF) IIB-like factor compound the fundamental core of all eukaryotic initiation complexes. The reason for the emergence and strict requirement of the additional intiation factor Bdp1, which is unique to the RNA polymerase (RNAP) III sytem, however, remained elusive. A poorly studied aspect in this context is the effect of DNA strain, that arises from DNA compaction and transcriptional activity, on the efficiency of initiation complex formation. We made use of a new nanotechnological tool – a DNA origami-based force clamp - to follow the assembly of human initiation complexes in the Pol II and Pol III system at the single-molecule level under piconewton forces. We demonstrate that TBP-DNA complexes are force-sensitive and TFIIB is necessary and sufficient to stabilise TBP on a strained RNAP II promoter. In contrast, Bdp1 is the pivotal component that ensures stable anchoring of initiation factors, and thus the polymerase itself, in the RNAP III system. Thereby, we offer an explanation for the crucial role of Bdp1 for the high transcriptional output of Pol III genes for the first time.

scholarly journals Multiplexed direct detection of barcoded protein reporters on a nanopore array

2019 ◽  
Author(s):  
Nicolas Cardozo ◽  
Karen Zhang ◽  
Katie Doroschak ◽  
Aerilynn Nguyen ◽  
Zoheb Siddiqui ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Direct Detection ◽  
Biological Activities ◽  
Sensor Technology ◽  
One Pot ◽  
Single Molecule Level ◽  
Dna And Rna ◽  
Oxford Nanopore ◽  
Reporter Proteins

AbstractGenetically encoded reporter proteins are a cornerstone of molecular biology. While they are widely used to measure many biological activities, the current number of uniquely addressable reporters that can be used together for one-pot multiplexed tracking is small due to overlapping detection channels such as fluorescence. To address this, we built an expanded library of orthogonally-barcoded Nanopore-addressable protein Tags Engineered as Reporters (NanoporeTERs), which can be read and demuxed by nanopore sensors at the single-molecule level. By adapting a commercially available nanopore sensor array platform typically used for real-time DNA and RNA sequencing (Oxford Nanopore Technologies’ MinION), we show direct detection of NanoporeTER expression levels from unprocessed bacterial culture with no specialized sample preparation. These results lay the foundations for a new class of reporter proteins to enable multiplexed, real-time tracking of gene expression with nascent nanopore sensor technology.

scholarly journals Local, Single-Molecule Oxidative Cleaving of DNA Origami by C60 on an AFM Tip

2021 ◽  
Author(s):  
Ankita Ray ◽  
Cristiana Passiu ◽  
S.N Ramakrishna ◽  
Antonella Rossi ◽  
Akinori Kuzuya ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Oxidative Damage ◽  
Single Molecule ◽  
Dna Cleavage ◽  
Dna Origami ◽  
Oxygen Species ◽  
Mica Surface ◽  
Single Molecule Level ◽  
Afm Tips ◽  
Morphology Changes

Spatially controlled single-molecule oxidation of DNA was performed by photocatalytic generation of singlet oxygen on chemically functionalized AFM tips. A waffle-type DNA origami deposited on a mica surface is site-specifically destroyed by generation of reactive oxygen species at the AFM tip, which was modified with C<sub>60</sub>-tripod photocatalyst. Upon AFM scanning under photoirradiation, DNA morphology changes, corresponding to oxidative damage were clearly observed at the single-molecule level. The DNA cleavage occurred with strict dependence on photoirradiation and the presence of C<sub>60 </sub>on the AFM tip.

scholarly journals Strong plasmonic enhancement of single molecule photostability in silver dimer optical antennas

Nanophotonics ◽  
2018 ◽  
Vol 7 (3) ◽  
pp. 643-649 ◽  
Author(s):  
Izabela Kaminska ◽  
Carolin Vietz ◽  
Álvaro Cuartero-González ◽  
Philip Tinnefeld ◽  
Antonio I. Fernández-Domínguez ◽  
...  
Keyword(s):  
Numerical Simulations ◽  
Single Molecule ◽  
Dna Origami ◽  
Plasmonic Nanoparticles ◽  
Optical Antennas ◽  
Plasmonic Enhancement ◽  
Rate Enhancement ◽  
Single Molecule Level ◽  
Radiative Rate

AbstractPhotobleaching is an effect terminating the photon output of fluorophores, limiting the duration of fluorescence-based experiments. Plasmonic nanoparticles (NPs) can increase the overall fluorophore photostability through an enhancement of the radiative rate. In this work, we use the DNA origami technique to arrange a single fluorophore in the 12-nm gap of a silver NP dimer and study the number of emitted photons at the single molecule level. Our findings yielded a 30× enhancement in the average number of photons emitted before photobleaching. Numerical simulations are employed to rationalize our results. They reveal the effect of silver oxidation on decreasing the radiative rate enhancement.

Sign in / Sign up

Export Citation Format

Share Document