Search

Simulations of nucleic acids at different levels of structural details are increasingly used to complement and interpret experiments in different fields, from biophysics to medicine and materials science. However, the various structural models currently available for DNA and RNA and their accompanying suites of computational tools can be very rarely used in a synergistic fashion. The tacoxDNA webserver and standalone software package presented here are a step toward a long-sought interoperability of nucleic acids models. The webserver offers a simple interface for converting various common input formats of DNA structures and setting up molecular dynamics (MD) simulations. Users can, for instance, design DNA rings with different topologies, such as knots, with and without supercoiling, by simply providing an XYZ coordinate file of the DNA centre-line. More complex DNA geometries, as designable in the cadnano, CanDo and Tiamat tools, can also be converted to all-atom or oxDNA representations, which can then be used to run MD simulations. Though the latter are currently geared toward the native and LAMMPS oxDNA representations, the open-source package is designed to be further expandable. TacoxDNA is available at http://tacoxdna.sissa.it. © 2019 Wiley Periodicals, Inc., (© 2019 Wiley Periodicals, Inc.)

Knots and supercoiling are both introduced in bacterial plasmids by catalytic processes involving DNA strand passages. While the effects on plasmid organization has been extensively studied for knotting and supercoiling taken separately, much less is known about their concurrent action. Here, we use molecular dynamics simulations and oxDNA, an accurate mesoscopic DNA model, to study the kinetic and metric changes introduced by complex (five-crossing) knots and supercoiling in 2 kbp-long DNA rings. We find several unexpected results. First, the conformational ensemble is dominated by two distinct states, differing in branchedness and knot size. Secondly, fluctuations between these states are as fast as the metric relaxation of unknotted rings. In spite of this, certain boundaries of knotted and plectonemically-wound regions can persist over much longer timescales. These pinned regions involve multiple strands that are interlocked by the cooperative action of topological and supercoiling constraints. Their long-lived character may be relevant for the simplifying action of topoisomerases.

In living cells, DNA is highly confined in space with the help of condensing agents, DNA binding proteins and high levels of supercoiling. Due to challenges associated with experimentally studying DNA under confinement, little is known about the impact of spatial confinement on the local structure of the DNA. Here, we have used well characterized slits of different sizes to collect high resolution atomic force microscopy images of confined circular DNA with the aim of assessing the impact of the spatial confinement on global and local conformational properties of DNA. Our findings, supported by numerical simulations, indicate that confinement imposes a large mechanical stress on the DNA as evidenced by a pronounced anisotropy and tangent-tangent correlation function with respect to non-constrained DNA. For the strongest confinement we observed nanometer sized hairpins and interwound structures associated with the nicked sites in the DNA sequence. Based on these findings, we propose that spatial DNA confinement in vivo can promote the formation of localized defects at mechanically weak sites that could be co-opted for biological regulatory functions., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

We use an accurate coarse-grained model for DNA and stochastic molecular dynamics simulations to study the pore translocation of 10-kbp-long DNA rings that are knotted. By monitoring various topological and physical observables we find that there is not one, as previously assumed, but rather two qualitatively different modes of knot translocation. For both modes the pore obstruction caused by knot passage has a brief duration and typically occurs at a late translocation stage. Both effects are well in agreement with experiments and can be rationalized with a transparent model based on the concurrent tensioning and sliding of the translocating knotted chains. We also observed that the duration of the pore obstruction event is more controlled by the knot translocation velocity than the knot size. These features should advance the interpretation and design of future experiments aimed at probing the spontaneous knotting of biopolymers.

We use stochastic simulation techniques to sample the conformational space of linear semiflexible polymers in a crowded medium and study how the knotting properties depend on the crowder size and concentration. The abundance of physical knots in the chains, which for definiteness we model on 10 kb long DNA filaments, is shown to have a non-monotonic, unimodal dependence on the colloid diameter, dc. The maximum incidence of knots occurs when dc is about equal to half of the gyration radius of the isolated chain. The degree of enhancement of knots grows rapidly with the solution density and can be very conspicuous relative to the case of isolated chains with no crowders. For instance, at 30% volume fraction the relative increase is more than fourfold. This dramatic enhancement is shown to originate from the depletion-induced chain compaction over multiple and concurring length scales. The same effect accounts for the variations of the knot length that accompany the changes in knotting probability. The findings suggest that crowded media could be viably used as a passive physical means for controlling and modulating the incidence and length of knots in DNA and other types of semiflexible polymers.

Langevin dynamics simulations are used to characterize the typical mechanisms governing the spontaneous tying, untying and the dynamical evolution of knots in coarse-grained models of DNA chains confined in nanochannels. In particular we focus on how these mechanisms depend on the chain contour length, Lc, at a fixed channel width D = 56 nm corresponding to the onset of the Odijk scaling regime where chain backfoldings and hence knots are disfavoured but not suppressed altogether. We find that the lifetime of knots grows significantly with Lc, while that of unknots varies to a lesser extent. The underlying kinetic mechanisms are clarified by analysing the evolution of the knot position along the chain. At the considered confinement, in fact, knots are typically tied by local backfoldings of the chain termini where they are eventually untied after a stochastic motion along the chain. Consequently, the lifetime of unknots is mostly controlled by backfoldings events at the chain ends, which is largely independent of Lc. The lifetime of knots, instead, increases significantly with Lc because knots can, on average, travel farther along the chain before being untied. The observed interplay of knots and unknots lifetimes underpins the growth of the equilibrium knotting probability of longer and longer chains at fixed channel confinement.

The dynamical properties of entangled polyelectrolytes are investigated theoretically and computationally for a proposed novel micromanipulation setup. Specifically, we investigate the effects of DC and AC electric fields acting longitudinally on knotted DNA chains, modelled as semiflexible chains of charged beads, under mechanical tension. We consider various experimentally accessible values of the field amplitude and frequency as well as several of the simplest knot types. In particular, we consider both torus and twist knots because they are respectively known to be able or unable to slide along macroscopic threads and ropes. Strikingly, this qualitative distinction disappears in this microscopic context because all the considered knot types acquire a systematic drift in the direction of the electric force. Notably, the knot drift velocity and diffusion coefficient in zero field (both measurable also experimentally) can be used to define a characteristic "frictional" lengthscale for the various knot types. This previously unexplored length provides valuable information on the extent of self-interactions in the nominal knotted region. It is finally observed that the motion of a knot can effectively follow the AC field only if the driving period is larger than the knot relaxation time (for which the self-diffusion time provides an upper bound). These results suggest that salient aspects of the intrinsic dynamics of knots in DNA chains could be probed experimentally by means of external, time-dependent electric fields.

The amount and type of self-entanglement of DNA filaments is significantly affected by spatial confinement, which is ubiquitous in biological systems. Motivated by recent advancements in single DNA molecule experiments based on nanofluidic devices and by the introduction of algorithms capable of detecting knots in open chains, we investigate numerically the entanglement of linear, open DNA chains confined inside nano-slits. The results regard the abundance, type, and length of occurring knots and are compared with recent findings for DNA inside nano-channels. In both cases, the width of the confining region, D, spans the 30 nm-1 μm range and the confined DNA chains are 1-4 μm long. It is found that the knotting probability is maximum for slit widths in the 70-100 nm range. However, over the considered DNA contour lengths, the maximum incidence of knots remains below 20%, while for channel confinement it tops 50%. Further differences of the entanglement are seen for the average contour length of the knotted region, which drops significantly below D ~100 nm for channel-confinement, while it stays approximately constant for slit-like confinement. These properties ought to reverberate in different kinetic properties of linear DNA depending on confinement and could be detectable experimentally or exploitable in nano-technological applications.

Nanografted monolayers (NAMs) of DNA show novel physico-chemical properties that make them ideally suited for advanced biosensing applications. In comparison with alternative solid-phase techniques for diagnostic DNA detection, NAMs have the advantage of combining a small size with a high homogeneity of the DNA surface coverage. These two properties favour the extreme miniaturization and ultrasensitivity in high-throughput biosensing devices. The systematic use of NAMs for quantitative DNA (and protein) detection has so far suffered from the lack of a control on key fabrication parameters, such as the ss- or ds-DNA surface coverage. Here we report on a combined experimental-computational study that allows us to estimate the surface density of the grafted DNA by analyzing the sample mechanical response, that is the DNA patch height vs. applied tip load curves. It is shown that the same analysis scheme can be used to detect the occurrence of hybridization with complementary strands in solution and estimate its efficiency. Thanks to these quantitative relationships it is possible to use a single AFM-based setup to: (i) fabricate a DNA NAM, (ii) control the DNA surface coverage, and (iii) characterize its level of hybridization helping the design of NAMs with pre-determined fabrication parameters.

DNA and RNA polymerases active on bacterial and human genomes in the crowded environment of a cell are modeled as beads spaced along a string. Aggregation of the large polymerizing complexes increases the entropy of the system through an increase in entropy of the many small crowding molecules; this occurs despite the entropic costs of looping the intervening DNA. Results of a quantitative cost/benefit analysis are consistent with observations that active polymerases cluster into replication and transcription "factories" in both pro- and eukaryotes. We conclude that the second law of thermodynamics acts through nonspecific entropic forces between engaged polymerases to drive the self-organization of genomes into loops containing several thousands (and sometimes millions) of basepairs.

We present a numerical characterization of the statics and dynamics of the packaging of a semi-flexible polymer inside a sphere. The study is motivated by recent experiments on the packaging of DNA inside viral capsids. It is found that the force required to confine the coarse-grained polymer is in fair agreement with that found in experiments for the packaging of the phi29 bacteriophage genome. Despite its schematic nature, the model is capable of reproducing the most salient dynamical features of packaging experiments such as the presence of pauses during individual packaging processes and the trend of the resisting force as a function of chain packed fraction. [ABSTRACT FROM AUTHOR]