The Effects of Base Stacking on DNA Flexibility

Abstract

DNA is the main genetic component of life and understanding its basic properties is crucial for advancement in applied sciences. DNA, a long macromolecule, must complex with proteins and other molecules for packaging and condensing to fit in the nucleus of a cell. Persistence length is a physical property that defines the stiffness of a polymer, ranging from rod-like to string-like. The persistence length of double-stranded DNA is ∼150bp or ∼50nm making it a relatively inflexible molecule. It is hypothesized that the two main contributors limiting DNA flexibility are mutual charge repulsions along the DNA backbones and attractive base stacking interactions. However, the relative contributions to DNA stiffness of electrostatic repulsion and base stacking forces are unknown. We use experimental T4 DNA ligase-mediated cyclization kinetics experiments to measure the physical properties of DNA, specifically DNA longitudinal and torsional flexibilities and helical repeat.We measured the impact on DNA stiffness of enhancing or diminishing stacking energy involving modified adenine and guanine bases. Diaminopurine (DAP) is hypothesized to increase DNA bending stiffness by changing the base geometry and dipole moment so as to enhance stacking energy and inosine (dITP) is hypothesized to decrease DNA bending stiffness. Cyclization assays were performed with radiolabeled DNA probes varying in length from 211bp to 201bp. The free ends of the probes are complementary and, in the presence of T4 DNA ligase, yield various linear and circular species in vitro. The J-factor (related to persistence length through the worm-like chain polymer model) can be estimated in such assays. We found that the bending persistence length changed slightly relative to natural DNA but it is the twist persistence length that has a more dramatic change.