Site-specific free energy surface parameters from single-molecule fluorescence measurements of exciton-coupled (iCy3)2 dimer probes positioned at DNA replication fork junctions

Jack Maurer ^1,2,3, Claire S Albrecht ^1,2,4, Peter H von Hippel ^2,3

⁰Center for Optical, Molecular and Quantum Science, University of Oregon, Eugene, OR 97403, USA.

Nucleic Acids Research +

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February 5, 2025

View abstract on PubMed

Summary

DNA breathing, or local fluctuations, at single-stranded-double-stranded DNA junctions influences protein interactions. This study quantifies DNA breathing dynamics, revealing key molecular insights for DNA repair and replication proteins.

Area Of Science

Molecular Biology
Biophysics
Structural Biology

Background

DNA replication, recombination, and repair involve proteins recognizing specific DNA structures like replication forks and primer-template junctions.
DNA 'breathing' refers to local, thermally induced fluctuations in DNA structure, potentially influencing protein binding and function at critical junctions.

Purpose Of The Study

To theoretically analyze recent experimental data on DNA breathing dynamics at ss-dsDNA junctions.
To quantify the stability and energy barriers of DNA breathing events at specific sites within these junctions.

Main Methods

Theoretical analysis of polarization-sweep single-molecule fluorescence (PS-SMF) data.
Modeling of exciton-coupled dimer-labeled ss-dsDNA fork constructs.

Main Results

Identified four quasi-stable conformational macrostates associated with DNA breathing near junctions.
Quantified the relative stabilities and activation energy barriers for these DNA breathing events.
Demonstrated that DNA breathing dynamics are dependent on the probe's position relative to the junction.

Conclusions

Provided a detailed molecular model for DNA breathing at ss-dsDNA junctions.
Offered insights into the molecular mechanisms of proteins interacting with these DNA structures.
Highlighted the functional significance of DNA breathing in DNA metabolism processes.