Atomic Force Microscopy of DNA and DNA-Protein Interactions

Haynes, Philip J.; Main, Kavit; Akpinar, Bernice; Pyne, Alice L. B.

doi:10.1007/978-1-0716-2221-6_5

Cited by 8 publications

(4 citation statements)

References 31 publications

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“…3). This differed to previous experiments where nickel chloride was used to deposit DNA for AFM analysis 64 .…”

Section: Resultsmentioning

confidence: 62%

Under or Over? Tracing Complex DNA Structures with High Resolution Atomic Force Microscopy

Holmes,

Gamill,

Provan

et al. 2024

Preprint

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The intricate topology of DNA plays a crucial role in the regulation of cellular processes and genome stability1–4. Despite its significance, DNA topology is challenging to explicitly determine due to the length and conformational complexity of individual topologically constrained DNA molecules. Here, we introduce an innovative approach combining high-resolution Atomic Force Microscopy (AFM) imaging with automated computational analysis to directly determine DNA topology. Our pipeline enables high-throughput tracing of uncoated circular DNA molecules directly in aqueous conditions, enabling determination of the order of DNA crossings, i.e. which molecule passes over which. By accurately tracing the DNA path through every DNA crossing, we can explicitly determine DNA topology and precisely classify knots and catenanes. We validate our method using known catenated products of theE. coliXer recombination system and confirm the predicted topology of knotted products from this system. Our study uncovers a recurrent depositional effect for the Xer catenanes and uncovers the origins of this effect using coarse-grained simulations, and enhances the statistical robustness. Our pipeline is applicable to other DNA/RNA-protein structures, particularly those with inherent flexibility, and opens up avenues for understanding fundamental biological processes which are regulated by or affect DNA topology.

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“…3). This differed to previous experiments where nickel chloride was used to deposit DNA for AFM analysis 64 .…”

Section: Resultsmentioning

confidence: 62%

Under or Over? Tracing Complex DNA Structures with High Resolution Atomic Force Microscopy

Holmes,

Gamill,

Provan

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…In recent years, AFM has been used to study complex biological processes like protein aggregation and folding, membrane protein dynamics, and motor protein movement. − AFM imaging has allowed researchers to visualize DNA supersecondary structures and DNA–protein interactions including helicases as well. , The interaction of various DNA binding proteins like RecG and SSB have already been investigated by using AFM. In a recent study using AFM, Sun et al have shown that the loading of RecG helicase on the fork-DNA is facilitated by the single-strand binding protein (SSB) and the interaction between RecG and SSB leads to RecG remodeling, and as a result, RecG gets separated from the fork but remains bound to the duplex DNA.…”

Section: Atomic Force Microscopymentioning

confidence: 99%

Insights into the Dynamics and Helicase Activity of RecD2 of Deinococcus radiodurans during DNA Repair: A Single-Molecule Perspective

2023

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While the double helix is the most stable conformation of DNA inside cells, its transient unwinding and subsequent partial separation of the two complementary strands yields an intermediate single-stranded DNA (ssDNA). The ssDNA is involved in all major DNA transactions such as replication, transcription, recombination, and repair. The process of DNA unwinding and translocation is shouldered by helicases that transduce the chemical energy derived from nucleotide triphosphate (NTP) hydrolysis to mechanical energy and utilize it to destabilize hydrogen bonds between complementary base pairs. Consequently, a comprehensive understanding of the molecular mechanisms of these enzymes is essential. In the last few decades, a combination of single-molecule techniques (force-based manipulation and visualization) have been employed to study helicase action. These approaches have allowed researchers to study the single helicase−DNA complex in real-time and the free energy landscape of their interaction together with the detection of conformational intermediates and molecular heterogeneity during the course of helicase action. Furthermore, the unique ability of these techniques to resolve helicase motion at nanometer and millisecond spatial and temporal resolutions, respectively, provided surprising insights into their mechanism of action. This perspective outlines the contribution of single-molecule methods in deciphering molecular details of helicase activities. It also exemplifies how each technique was or can be used to study the helicase action of RecD2 in recombination DNA repair.

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“…experimental techniques, such as cryo-electron microscopy, X-ray crystallography, NMR spectroscopy, and other single-molecule techniques (e.g., light/magnetic tweezers and atomic force microscopy) [15][16][17][18]. These experimental methods face difficulties in elucidating the underlying aspects of DNA folding, hybridization, and stability.…”

Section: Introductionmentioning

confidence: 99%

“…Therefore, understanding the 3D structures and properties of DNA (e.g., dynamics, thermodynamics, and mechanics) is useful in understanding its biological functions and designing DNA nanomaterials [ 2 , 3 , 4 , 5 , 6 , 14 ]. However, the flexibility and polymorphism of DNA present challenges for current experimental techniques, such as cryo-electron microscopy, X-ray crystallography, NMR spectroscopy, and other single-molecule techniques (e.g., light/magnetic tweezers and atomic force microscopy) [ 15 , 16 , 17 , 18 ]. These experimental methods face difficulties in elucidating the underlying aspects of DNA folding, hybridization, and stability.…”

Section: Introductionmentioning

confidence: 99%

Computational Modeling of DNA 3D Structures: From Dynamics and Mechanics to Folding

Tan

Liu

et al. 2023

Molecules

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DNA carries the genetic information required for the synthesis of RNA and proteins and plays an important role in many processes of biological development. Understanding the three-dimensional (3D) structures and dynamics of DNA is crucial for understanding their biological functions and guiding the development of novel materials. In this review, we discuss the recent advancements in computer methods for studying DNA 3D structures. This includes molecular dynamics simulations to analyze DNA dynamics, flexibility, and ion binding. We also explore various coarse-grained models used for DNA structure prediction or folding, along with fragment assembly methods for constructing DNA 3D structures. Furthermore, we also discuss the advantages and disadvantages of these methods and highlight their differences.

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Atomic Force Microscopy of DNA and DNA-Protein Interactions

Cited by 8 publications

References 31 publications

Under or Over? Tracing Complex DNA Structures with High Resolution Atomic Force Microscopy

Under or Over? Tracing Complex DNA Structures with High Resolution Atomic Force Microscopy

Insights into the Dynamics and Helicase Activity of RecD2 of Deinococcus radiodurans during DNA Repair: A Single-Molecule Perspective

Computational Modeling of DNA 3D Structures: From Dynamics and Mechanics to Folding

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