Mechanical Analysis Methodology for DNA Minicircles Observed by Cryo-Em

Abstract

Recent cryo-electron microscopy (cryo-EM) images of DNA minicircles, about 100 basepairs in length, provide a new perspective on the mechanics of DNA. In essence, the resultant 3-dimensional reconstructions capture the mechanically deformed state of the double helix at an instant in time. Such deformations may include sites with high bending and/or torsional flexibility that could result from the tight bending required to cyclize, superhelical stress, and thermal fluctuations. Unfortunately, these reconstructions resolve only the DNA helical axis and provide no information about (i) how to register their known basepair sequence with the reconstruction and (ii) how torsional deformations are distributed along the length of the minicircle. In addition, the experimental procedures are complicated and consequently limit the the number of reconstructed minicircles to about 20. Our objective is to understand the mechanics of these DNA minicircles, and specifically, to develop a method capable of detecting the presence of kinks or torsional destabilizations in their cryo-EM reconstructions. To this end, we developed a modal analysis approach to describe the mechanics of DNA minicircles. In our method, we use the thermal modes of a homogeneous elastic rod representation for circular DNA to assign modal amplitudes to each reconstruction. The distribution of modal amplitudes for a population of minicircles provides a unique ‘signature’ dependent upon several variables, including superhelical density and the presence of kinks or torsional destabilizations. To test our method and predict these signatures, we developed a statistical mechanics model to simulate ensembles of DNA minicircles. This model can represent sequence dependence (elasticity/curvature) and prescribed kinks or torsional destabilizations. Our preliminary analysis suggests that the observed signatures are inconsistent with a homogeneous elastic rod and thereby implicate the role of sequence dependence, kinks or torsional instabilities.