Real-Time Imaging of Platinum−Sulfur Ligand Exchange Reactions at the Single-Molecule Level via a General Chemical Technique

2011 ◽  
Vol 30 (11) ◽  
pp. 2901-2907 ◽  
Author(s):  
N. Melody Esfandiari ◽  
Yong Wang ◽  
Theresa M. McIntire ◽  
Suzanne A. Blum
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Ligand Exchange ◽  
Exchange Reactions ◽  
Real Time Imaging ◽  
Single Molecule Level ◽  
General Chemical ◽  
Chemical Technique ◽  
Sulfur Ligand

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

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 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 Real-time tracking of single-molecule collagenase on native collagen and partially structured collagen-mimic substrates

2018 ◽  
Vol 54 (73) ◽  
pp. 10248-10251
Author(s):  
James Froberg ◽  
Woo-Sik Choi ◽  
Abbas Sedigh ◽  
Tayebeh Anajafi ◽  
Jasmin Farmakes ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Real Time Imaging ◽  
Native Collagen ◽  
Proteolytic Activities ◽  
Real Time Tracking

Real-time imaging and tracking of proteolytic activities of individual enzymes with their native and structurally modified substrates has been investigated.

scholarly journals A label-free approach to detect ligand binding to cell surface proteins in real time

2018 ◽  
Author(s):  
Verena Burtscher ◽  
Matej Hotka ◽  
Yang Li ◽  
Michael Freissmuth ◽  
Walter Sandtner
Keyword(s):  
Ligand Binding ◽  
Real Time ◽  
Single Molecule ◽  
Membrane Capacitance ◽  
Surface Proteins ◽  
Displacement Current ◽  
High Temporal Resolution ◽  
Label Free ◽  
Single Molecule Level ◽  
Electrophysiological Recordings

AbstractElectrophysiological recordings allow for monitoring the operation of proteins with high temporal resolution down to the single molecule level. This technique has been exploited to track either ion flow arising from channel opening or the synchronized movement of charged residues and/or ions within the membrane electric field. Here, we describe a novel type of current by using the serotonin transporter (SERT) as a model. We examined transient currents elicited on rapid application of specific SERT inhibitors. Our analysis shows that these currents originate from ligand binding and not from a conformational change. The Gouy-Chapman model predicts that a ligand-induced elimination/neutralization of surface charge must produce a displacement current and related apparent changes in membrane capacitance. Here we verified these predictions with SERT. Our observations demonstrate that ligand binding to a protein can be monitored in real time and in a label-free manner by recording the membrane capacitance.

scholarly journals Active DNA unwinding and transport by a membrane-adapted helicase nanopore

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Ke Sun ◽  
Changjian Zhao ◽  
Xiaojun Zeng ◽  
Yuejia Chen ◽  
Xin Jiang ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Single Cell Analysis ◽  
Single Channel ◽  
Vital Role ◽  
Live Cell ◽  
Bovine Papillomavirus ◽  
Nanoscale Transport ◽  
Single Molecule Level

Abstract Nanoscale transport through nanopores and live-cell membranes plays a vital role in both key biological processes as well as biosensing and DNA sequencing. Active translocation of DNA through these nanopores usually needs enzyme assistance. Here we present a nanopore derived from truncated helicase E1 of bovine papillomavirus (BPV) with a lumen diameter of c.a. 1.3 nm. Cryogenic electron microscopy (cryo-EM) imaging and single channel recording confirm its insertion into planar lipid bilayer (BLM). The helicase nanopore in BLM allows the passive single-stranded DNA (ssDNA) transport and retains the helicase activity in vitro. Furthermore, we incorporate this helicase nanopore into the live cell membrane of HEK293T cells, and monitor the ssDNA delivery into the cell real-time at single molecule level. This type of nanopore is expected to provide an interesting tool to study the biophysics of biomotors in vitro, with potential applications in biosensing, drug delivery and real-time single cell analysis.

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