Relative Contributions of Base Stacking and Electrostatic Repulsion on DNA Nicks and Gaps
KAUST DepartmentBiological and Environmental Sciences and Engineering (BESE) Division
Laboratory of DNA Replication and Recombination
Single-Molecule Spectroscopy and Microscopy Research Group
KAUST Grant NumberUFR/1/3764-01-01
Embargo End Date2021-11-12
Permanent link to this recordhttp://hdl.handle.net/10754/665998
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AbstractIn duplex DNA, the continuous sugar phosphate backbones prevent the double helix from significant bending, but breaks in the duplex such as nicks, gaps, and flaps present points at which significant bending is possible. The conformational dynamics of these aberrant structures remains poorly understood. Two factors can maintain the duplexlike conformation of these aberrant structures, these being the hydrophobic and aromatic stacking interactions of the nucleobases, and the electrostatic repulsion of the negatively charged backbones. Using confocal single-molecule Förster resonance energy transfer on nicked and gapped DNA structures, we compare the relative contributions of these two factors by modulating the electrostatic repulsion through mono- and divalent cation concentrations. Base stacking interactions dominate the dynamics of nicked DNA, making it behave essentially like duplex DNA. Gapped structures have weaker base stacking and thus backbone electrostatic repulsion becomes important, and shielding from cations results in an average increase in bending around the gap. This bending of gapped structures could be interpreted by increased flexibility of unstacked structures, transient unstacking events, or a combination of the two. Burst variance analysis (BVA) and analysis by photon-by-photon hidden Markov modeling (H2MM), methods capable of detecting submillisecond dynamics of single molecules in solution, only revealed a single state, indicating that dynamics are occurring at time scales shorter than microseconds.
CitationHarris, P. D., Hamdan, S. M., & Habuchi, S. (2020). Relative Contributions of Base Stacking and Electrostatic Repulsion on DNA Nicks and Gaps. The Journal of Physical Chemistry B. doi:10.1021/acs.jpcb.0c06941
SponsorsThe authors thank Dr. Eitan Lerner for his invaluable help using FRETBursts, for writing the Jupyter notebooks used to analyze the data, and for consultation on best background thresholds and H2 MM parameters. We also thank Vlad-Stefan Raducanu for his consultation regarding labeling procedures and data interpretation. The research reported in this publication was supported by King Abdullah University of Science and Technology (KAUST), and the KAUST Office of Sponsored Research (OSR) under Award No. UFR/1/3764-01-01.
PublisherAmerican Chemical Society (ACS)
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