The Impact of Base Stacking on the Conformations and Electrostatics of Single-Stranded DNA

Single-stranded DNA (ssDNA) is notable for its interactions with ssDNA binding proteins (SSBs) during fundamentally important biological processes including DNA repair and replication. Previous work has begun to characterize the conformational and electrostatic properties of ssDNA in association with SSBs. However, the conformational distributions of free ssDNA have been difficult to determine. To capture the vast array of ssDNA conformations in solution, we pair small angle X-ray scattering with novel ensemble fitting methods, obtaining key parameters such as the size, shape and stacking character of strands with different sequences. Complementary ion counting measurements using inductively coupled plasma atomic emission spectroscopy are employed to determine the composition of the ion atmosphere at physiological ionic strength. Applying this combined approach to poly dA and poly dT, we find that the global properties of these sequences are very similar, despite having vastly different propensities for single-stranded helical stacking. These results suggest that a relatively simple mechanism for the binding of ssDNA to non-specific SSBs may be at play, which explains the disparity in binding affinities observed for these systems

Complexities in barrier island response to sea level rise : insights from numerical model experiments, North Carolina Outer Banks

Author Posting. © American Geophysical Union, 2010. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research 115 (2010): F03004, doi:10.1029/2009JF001299.Using a morphological-behavior model to conduct sensitivity experiments, we investigate the sea level rise response of a complex coastal environment to changes in a variety of factors. Experiments reveal that substrate composition, followed in rank order by substrate slope, sea level rise rate, and sediment supply rate, are the most important factors in determining barrier island response to sea level rise. We find that geomorphic threshold crossing, defined as a change in state (e.g., from landward migrating to drowning) that is irreversible over decadal to millennial time scales, is most likely to occur in muddy coastal systems where the combination of substrate composition, depth-dependent limitations on shoreface response rates, and substrate erodibility may prevent sand from being liberated rapidly enough, or in sufficient quantity, to maintain a subaerial barrier. Analyses indicate that factors affecting sediment availability such as low substrate sand proportions and high sediment loss rates cause a barrier to migrate landward along a trajectory having a lower slope than average barrier island slope, thereby defining an “effective” barrier island slope. Other factors being equal, such barriers will tend to be smaller and associated with a more deeply incised shoreface, thereby requiring less migration per sea level rise increment to liberate sufficient sand to maintain subaerial exposure than larger, less incised barriers. As a result, the evolution of larger/less incised barriers is more likely to be limited by shoreface erosion rates or substrate erodibility making them more prone to disintegration related to increasing sea level rise rates than smaller/more incised barriers. Thus, the small/deeply incised North Carolina barriers are likely to persist in the near term (although their long-term fate is less certain because of the low substrate slopes that will soon be encountered). In aggregate, results point to the importance of system history (e.g., previous slopes, sediment budgets, etc.) in determining migration trajectories and therefore how a barrier island will respond to sea level rise. Although simple analytical calculations may predict barrier response in simplified coastal environments (e.g., constant slope, constant sea level rise rate, etc.), our model experiments demonstrate that morphological-behavior modeling is necessary to provide critical insights regarding changes that may occur in environments having complex geometries, especially when multiple parameters change simultaneously.This work was partially supported by the U.S. Geological Survey, Woods Hole Science Center and a sabbatical leave fellowship from Oberlin College to Laura Moore from the Mellon‐8 Consortium

Progress in small-angle scattering from biological solutions at high-brilliance synchrotrons

Small-angle X-ray scattering (SAXS) is an established technique that provides low-resolution structural information on macromolecular solutions. Recent decades have witnessed significant progress in both experimental facilities and in novel data-analysis approaches, making SAXS a mainstream method for structural biology. The technique is routinely applied to directly reconstruct low-resolution shapes of proteins and to generate atomistic models of macromolecular assemblies using hybrid approaches. Very importantly, SAXS is capable of yielding structural information on systems with size and conformational polydispersity, including highly flexible objects. In addition, utilizing high-flux synchrotron facilities, time-resolved SAXS allows analysis of kinetic processes over time ranges from microseconds to hours. Dedicated bioSAXS beamlines now offer fully automated data-collection and analysis pipelines, where analysis and modelling is conducted on the fly. This enables SAXS to be employed as a high-throughput method to rapidly screen various sample conditions and additives. The growing SAXS user community is supported by developments in data and model archiving and quality criteria. This review illustrates the latest developments in SAXS, in particular highlighting time-resolved applications aimed at flexible and evolving systems

Both helix topology and counterion distribution contribute to the more effective charge screening in dsRNA compared with dsDNA

The recent discovery of the RNA interference mechanism emphasizes the biological importance of short, isolated, double-stranded (ds) RNA helices and calls for a complete understanding of the biophysical properties of dsRNA. However, most previous studies of the electrostatics of nucleic acid duplexes have focused on DNA. Here, we present a comparative investigation of electrostatic effects in RNA and DNA. Using resonant (anomalous) and non-resonant small-angle X-ray scattering, we characterized the charge screening efficiency and counterion distribution around short (25 bp) dsDNA and RNA molecules of comparable sequence. Consistent with theoretical predictions, we find counterion mediated screening to be more efficient for dsRNA than dsDNA. Furthermore, the topology of the RNA A-form helix alters the spatial distribution of counterions relative to B-form DNA. The experimental results reported here agree well with ion-size-corrected non-linear Poisson–Boltzmann calculations. We propose that differences in electrostatic properties aid in selective recognition of different types of short nucleic acid helices by target binding partners