Blind Predictions of DNA and RNA Tweezers Experiments with Force and Torque

Single-molecule tweezers measurements of double-stranded nucleic acids (dsDNA and dsRNA) provide unprecedented opportunities to dissect how these fundamental molecules respond to forces and torques analogous to those applied by topoisomerases, viral capsids, and other biological partners. However, tweezers data are still most commonly interpreted post facto in the framework of simple analytical models. Testing falsifiable predictions of state-of-the-art nucleic acid models would be more illuminating but has not been performed. Here we describe a blind challenge in which numerical predictions of nucleic acid mechanical properties were compared to experimental data obtained recently for dsRNA under applied force and torque. The predictions were enabled by the HelixMC package, first presented in this paper. HelixMC advances crystallography-derived base-pair level models (BPLMs) to simulate kilobase-length dsDNAs and dsRNAs under external forces and torques, including their global linking numbers. These calculations recovered the experimental bending persistence length of dsRNA within the error of the simulations and accurately predicted that dsRNA's “spring-like” conformation would give a two-fold decrease of stretch modulus relative to dsDNA. Further blind predictions of helix torsional properties, however, exposed inaccuracies in current BPLM theory, including three-fold discrepancies in torsional persistence length at the high force limit and the incorrect sign of dsRNA link-extension (twist-stretch) coupling. Beyond these experiments, HelixMC predicted that ‘nucleosome-excluding’ poly(A)/poly(T) is at least two-fold stiffer than random-sequence dsDNA in bending, stretching, and torsional behaviors; Z-DNA to be at least three-fold stiffer than random-sequence dsDNA, with a near-zero link-extension coupling; and non-negligible effects from base pair step correlations. We propose that experimentally testing these predictions should be powerful next steps for understanding the flexibility of dsDNA and dsRNA in sequence contexts and under mechanical stresses relevant to their biology.Author Summary: DNA and RNA are fundamental molecules in the central dogma of molecular biology. Many biological behaviors of double-stranded DNA and RNA – including transcription/translation by proteins and packaging into compact structures – depend on their ability to flex and twist. Single-molecule tweezers now provide accurate mechanical measurements of DNA and RNA helices under force and torque but have not been used to rigorously falsify and thereby advance computational models. Here we present the first such blind challenge, involving recent dsRNA tweezers data that were kept hidden from modelers and a new HelixMC toolkit that resolves challenges in simulating long double helices from base-pair level models. The predictions gave excellent agreement with bending and stretching measurements of dsRNA but failed to recover twisting properties, pinpointing a critical area of future investigation. HelixMC also predicted that poly(A)/poly(T) and Z-DNA–biologically important variants whose elastic responses have not been studied with tweezers–will have distinct mechanical properties. These results open a route to iteratively falsifying and refining computational models of long nucleic acid helices, as is necessary for attaining a predictive understanding of their biological behaviors.