Response of inhomogeneous DNA to external twist
Keywords: DNA damage
DNA nanotube
supercoiling

Studies of supercoiled DNA have typically treated DNA as a homogeneous polymer. In reality, however, DNA is highly inhomogeneous. Defects, for example, are an important source of inhomogeneities in DNA. Caused by processes that damage DNA, these defects can have strong effects on local elastic properties. In this thesis, we seek to address the question of how DNA supercoiling affects local defect properties, or, conversely, how the presence of such defects changes global supercoiled structure. Using a model of the thymine dimer (TD) —a major mutagenic photoproduct induced by UV radiation—in the oxDNA forcefield, we performed extensive molecular dynamics (MD) simulations of thymine dimer-containing double-stranded DNA (dsDNA) as a function of external twist and force. Compared to undamaged DNA, the presence of a thymine dimer lowers the supercoiling densities at which plectonemes and bubbles occur. For biologically relevant supercoiling densities and forces, thymine dimers are observed to preferentially segregate to the tips of the plectonemes, where they enhance the probability of a localized tip-bubble. This mechanism increases the probability of highly bent and denatured states at the thymine dimer site, which may facilitate repair enzyme binding. Thymine dimer-induced tip-bubbles are also observed to pin plectonemes, which may help repair enzymes to locate damage.
To understand the physics of how thymine dimers are segregated to the plectoneme tips in an extrahelical arrangement, we develop a free energy model of tip-bubble plectoneme formation. The model is capable of reproducing the major state boundaries in the force-torsion space of negatively supercoiled dsDNA, and predicting how variations in defect properties could affect the probability of plectoneme, bubble and tip-bubble formation in supercoiled dsDNA. The model is further validated by comparing its predictions with our MD simulation findings on thymine dimer-containing dsDNA, and stress-tested using MD simulations of dsDNA that contains other types of defects, such as base pair mismatches. We also expanded our analysis to the force-torsion space of positively supercoiled dsDNA, and showed that tip-bubbles formed in positively supercoiled dsDNA bear several physical characteristics that are different from its counterpart in negatively supercoiled dsDNA.
We also seek to address, by simulations, the fundamental question of how repair proteins search and identify defects in a crowded cellular environment. To do so, we modelled a crowded intranuclear environment in bacterial cells using the twistable elastic polymer (TEP) model, introduced a DNA defect model by reducing the bending stiffness of several DNA TEP particles, and constructed an actively rotating RNA polymerase model to generate supercoils. We observe that the DNA defect is localised at the tip of a plectonemic supercoil generated by the polymerase model away from the crowded intranuclear environment. This observation strengthens our hypothesis—originally proposed based on MD simulations of a simple molecular tweezer geometry—that the interaction between supercoiling and defects might facilitate damage recognition in living cells.
Finally, we extended our study of defects to DNA nanotubes. These nanotubes are rod-like nanostructures that comprise parallel arrangements of dsDNA. We performed twist simulations of DNA nanotubes similar to the ones we did for dsDNA and showed that the buckling of DNA nanotubes occurs in a defect-driven, non-wormlike chain regime. Our findings raise the question of whether theories based on the wormlike chain model can still be used to study the buckling transition and plectoneme formation in DNA nanotubes, as was done in several experimental studies.