Torque Spectroscopy of Sequence-Dependent Structural Transitions in DNA

Abstract

B-DNA becomes unstable under superhelical stress and is able to adopt a wide range of alternative conformations including strand-separated DNA and Z-DNA. Localized sequence-dependent structural transitions are important for the regulation of biological processes such as DNA replication and transcription. To directly probe the effect of sequence on structural transitions driven by torque, we have measured the torsional response of a panel of DNA sequences, using single molecule assays that employ nanosphere rotational probes to achieve high torque resolution. We have used AT-rich sequences to study strand separation, and d(pGpC)n sequences to study Z-DNA formation. For both types of sequences, we have observed responses that match our predictions based on a theoretical treatment of cooperative transitions in helical polymers. In the fixed twist ensemble, theory predicts that domain wall penalties should lead to significant torque overshoots, which are confirmed by our measurements. We have rigorously challenged our models by experimentally varying the boundary conditions, using atomic spacers introduced at the edges of sequences of interest. Our mechanical measurements include direct characterization of the torsional rigidities and helicities of non-canonical conformations, and establish a framework for quantitative predictions of the complex torsional response of arbitrary sequences in their biological context.