Investigation of DNA Sharp-Bending Defects through Pulling DNA Knots

Description

DNA sharp bending occurs in many biological processes, such as DNA wrapping around histones in nucleosomes. DNA sharp bending requires huge bending energy and how biological processes overcome the bending energy cost is not completely clear. In past years, the energy cost of DNA sharp bending was indirectly measured by a few experiments and the results suggested that the energy cost may be significantly less than the one estimated from the wormlike chain with the persistence length of ~50 nm. To explain these experimental observations, some researchers hypothesized that DNA becomes more flexible at sharp bending conditions due to the excitation of flexible defects, such as kinks and bubbles (local melting). Qualitatively, the hypothesis should be true: when the bending curvature exceeds a threshold, the DNA double-helix structure is disrupted, which leads to structural defect(s). Several key questions remain to be addressed: (i) what is the threshold curvature? (ii) what are the excitation energies of defects? (iii) what are the bending flexibilities of defects? The answers to these questions, i.e., quantitative properties of defects, determine whether and when DNA sharp-bending defects occur in biological processes. In this project, we will quantify the properties of DNA sharp-bending defects using a new approach: pulling DNA knots in single-DNA magnetic-tweezers experiments and multiscale simulations. As the pulling force increases, the knot core becomes smaller and the DNA bending curvature in the knot core increases. When the pulling force reaches a critical value, one or more DNA sharp-bending defects will be induced, and the DNA knot conformation undergoes equilibrium transition between the intact and defective states. The measurement of the critical pulling force and other quantities will be used to extract the critical bending curvature, defect excitation energy and defect flexibility. We will analyze the dependence of defect properties on temperature, salt concentration and DNA sequence. Pulling DNA knots will be carried out in multiscale simulations, including all-atom simulations, simulations based on the oxDNA model, coarse-grained simulations based on the bead-spring model. Single-DNA magnetic-tweezers experiments of pulling DNA knots will be carried out by our collaborator, who has successfully produced DNA knots in experiments. Our new approach, pulling DNA knots to induce sharp-bending defects, is likely to provide many new quantitative results of DNA sharp-bending defects from both experiments and simulations. The results should dramatically enhance our understanding of the biological processes involving DNA sharp bending.