scholarly journals Combining Single-molecule Manipulation and Imaging for the Study of Protein-DNA Interactions

10.3791/51446 ◽  
2014 ◽  
Author(s):  
Carina Monico ◽  
Gionata Belcastro ◽  
Francesco Vanzi ◽  
Francesco S. Pavone ◽  
Marco Capitanio
Keyword(s):  
Single Molecule ◽  
Dna Interactions ◽  
Protein Dna Interactions ◽  
Single Molecule Manipulation

scholarly journals Stretching and immobilization of DNA for studies of protein–DNA interactions at the single-molecule level

2007 ◽  
Vol 2 (4) ◽  
pp. 185-201 ◽  
Author(s):  
Ji Hoon Kim ◽  
Venkat Ram Dukkipati ◽  
Stella W. Pang ◽  
Ronald G. Larson
Keyword(s):  
Single Molecule ◽  
Dna Interactions ◽  
Single Molecule Level ◽  
Protein Dna Interactions

scholarly journals Shining a Spotlight on DNA: Single-Molecule Methods to Visualise DNA

Molecules ◽  
2019 ◽  
Vol 24 (3) ◽  
pp. 491 ◽  
Author(s):  
Gurleen Kaur ◽  
Jacob Lewis ◽  
Antoine van Oijen
Keyword(s):  
Mechanical Properties ◽  
Nucleic Acids ◽  
Small Molecules ◽  
Single Molecule ◽  
Single Molecules ◽  
Dna Molecules ◽  
Dna Interactions ◽  
Biochemical Reactions ◽  
Imaging Methods ◽  
Protein Dna Interactions

The ability to watch single molecules of DNA has revolutionised how we study biological transactions concerning nucleic acids. Many strategies have been developed to manipulate DNA molecules to investigate mechanical properties, dynamics and protein–DNA interactions. Imaging methods using small molecules and protein-based probes to visualise DNA have propelled our understanding of complex biochemical reactions involving DNA. This review focuses on summarising some of the methodological developments made to visualise individual DNA molecules and discusses how these probes have been used in single-molecule biophysical assays.

scholarly journals Engineered Holliday Junctions As Single-Molecule Reporters For Protein-Dna Interactions

2010 ◽  
Vol 98 (3) ◽  
pp. 74a-75a
Author(s):  
Nesha May O. Andoy ◽  
Susanta Sarkar ◽  
Peng Chen
Keyword(s):  
Single Molecule ◽  
Holliday Junctions ◽  
Dna Interactions ◽  
Protein Dna Interactions

scholarly journals Short-Read Single-Molecule DNA Sequencing for Highly Parallel Analysis of Protein-DNA Interactions

2018 ◽  
Vol 114 (3) ◽  
pp. 92a
Author(s):  
Rebecca Andrews ◽  
Horst Steuer ◽  
Arun Shivalingam ◽  
Afaf H. El-Sagheer ◽  
Tom Brown ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Single Molecule ◽  
Parallel Analysis ◽  
Dna Interactions ◽  
Short Read ◽  
Protein Dna Interactions

scholarly journals DNA Flow-Stretch Assays for Studies of Protein-DNA Interactions at the Single-Molecule Level

Applied Nano ◽  
2022 ◽  
Vol 3 (1) ◽  
pp. 16-41
Author(s):  
Aurimas Kopūstas ◽  
Mindaugas Zaremba ◽  
Marijonas Tutkus
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Dna Interaction ◽  
Dna Interactions ◽  
Ensemble Averaging ◽  
Target Search ◽  
Single Molecule Level ◽  
Protein Dna Interactions ◽  
Long Time

Protein-DNA interactions are the core of the cell’s molecular machinery. For a long time, conventional biochemical methods served as a powerful investigatory basis of protein-DNA interactions and target search mechanisms. Currently single-molecule (SM) techniques have emerged as a complementary tool for studying these interactions and have revealed plenty of previously obscured mechanistic details. In comparison to the traditional ones, SM methods allow direct monitoring of individual biomolecules. Therefore, SM methods reveal reactions that are otherwise hidden by the ensemble averaging observed in conventional bulk-type methods. SM biophysical techniques employing various nanobiotechnology methods for immobilization of studied molecules grant the possibility to monitor individual reaction trajectories of biomolecules. Next-generation in vitro SM biophysics approaches enabling high-throughput studies are characterized by much greater complexity than the ones developed previously. Currently, several high-throughput DNA flow-stretch assays have been published and have shown many benefits for mechanistic target search studies of various DNA-binding proteins, such as CRISPR-Cas, Argonaute, various ATP-fueled helicases and translocases, and others. This review focuses on SM techniques employing surface-immobilized and relatively long DNA molecules for studying protein-DNA interaction mechanisms.

Nanoscale “Curtain Rods”: High-Throughput Tools for Studying DNA-Protein Interactions

2008 ◽  
Vol 1138 ◽  
Author(s):  
Teresa Fazio ◽  
Mari-Liis Visnapuu ◽  
Shalom J. Wind ◽  
Eric Greene
Keyword(s):  
Data Acquisition ◽  
Real Time ◽  
Lipid Bilayer ◽  
High Throughput ◽  
Single Molecule ◽  
Protein Interactions ◽  
Massively Parallel ◽  
Dna Interactions ◽  
Parallel Data ◽  
Protein Dna Interactions

AbstractIn this work, we combine nanoscale engineering with single-molecule biology to probe the biochemical interactions between individual proteins and DNA. This approach, a vast improvement over previous methods, constructs a platform to observe thousands of protein-DNA interactions in real time with unprecedented detail. A key challenge in these experiments involves collecting enough statistically relevant data in order to analyze reactions which are designed to be probed individually. “DNA curtains” are formed by flowing the DNA tethered to a lipid bilayer across nanopatterned barriers, facilitating massively parallel data acquisition.

scholarly journals Homologous Recombination under the Single-Molecule Fluorescence Microscope

2019 ◽  
Vol 20 (23) ◽  
pp. 6102
Author(s):  
Dalton R. Gibbs ◽  
Soma Dhakal
Keyword(s):  
Homologous Recombination ◽  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Dna Interactions ◽  
Single Molecule Fluorescence ◽  
Spatiotemporal Resolution ◽  
Dynamic Interactions ◽  
Protein Dna Interactions ◽  
Dna Substrates ◽  
Single Molecule Fluorescence Microscopy

Homologous recombination (HR) is a complex biological process and is central to meiosis and for repair of DNA double-strand breaks. Although the HR process has been the subject of intensive study for more than three decades, the complex protein–protein and protein–DNA interactions during HR present a significant challenge for determining the molecular mechanism(s) of the process. This knowledge gap is largely because of the dynamic interactions between HR proteins and DNA which is difficult to capture by routine biochemical or structural biology methods. In recent years, single-molecule fluorescence microscopy has been a popular method in the field of HR to visualize these complex and dynamic interactions at high spatiotemporal resolution, revealing mechanistic insights of the process. In this review, we describe recent efforts that employ single-molecule fluorescence microscopy to investigate protein–protein and protein–DNA interactions operating on three key DNA-substrates: single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), and four-way DNA called Holliday junction (HJ). We also outline the technological advances and several key insights revealed by these studies in terms of protein assembly on these DNA substrates and highlight the foreseeable promise of single-molecule fluorescence microscopy in advancing our understanding of homologous recombination.

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