A Method for the Sensitive Detection of Base Pairing and Stacking Energies for Damaged and Noncanonical Sequences from Single Molecule Unfolding Experiments.

Micah J McCauley, Ioulia Rouzina, Megan E Núñez, Mark C Williams

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Article in Methods in molecular biology (Clifton, N.J.), 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

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4 authors.

Micah J McCauleyDepartment of Physics, Northeastern University, Boston, MA, USA.

Ioulia RouzinaDepartment of Chemistry and Biochemistry, The Ohio State University, Columbus, OH, USA.

Megan E NúñezDepartment of Chemistry and Program in Biochemistry, Wellesley College, Wellesley, MA, USA.

Mark C WilliamsDepartment of Physics, Northeastern University, Boston, MA, USA. ma.williams@northeastern.edu.

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6 · The paper itself

Abstract

Noncanonical and damaged bases occur with significant frequency and recognizing them is an important step in many nuclear processes including DNA repair, homologous recombination, and transcription. Yet while both the pairing and stacking energies of the canonical bases are well known in theory and experiment, the energetics of noncanonical bases are not well understood. Incorporating noncanonical bases into DNA hairpins affords the opportunity to quantify the changes in stacking and pairing interactions during forced hairpin unfolding. Though the measurement accuracy of this technique is quite high, technical challenges are significant and the overall throughput is challenging. In this chapter, we present a method for measuring the pairing and stacking energies of such noncanonical base pairs by incorporating the hairpin defect in the lower stem and performing optical tweezers force-unfolding experiments. During unfolding, this stem frays apart before the upper half, enhancing detection and facilitating rapid and sensitive characterization of these defects.

Indexed as

Base PairingDNADNA DamageSingle Molecule ImagingNucleic Acid ConformationOptical TweezersThermodynamicsDNADNADNA damageEnergy landscapeFree energyMfoldNucleic acidsOptical tweezersRNAUnfolding

Identifiers

PMID40175881

DOI10.1103/PhysRevLett.118.088101

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