Bridging the solution divide: comprehensive structural analyses of dynamic RNA, DNA, and protein assemblies by small-angle X-ray scattering

Abstract

Summary Small-Angle X-ray Scattering (SAXS) is changing how we perceive biological structures, because it reveals dynamic macromolecular conformations and assemblies in solution. SAXS information captures thermodynamic ensembles, enhances static structures detailed by high-resolution methods, uncovers commonalities among diverse macromolecules, and helps define biological mechanisms. SAXS-based experiments on RNA riboswitches and ribozymes and on DNA-protein complexes including DNA-PK and p53 discover flexibilities that better define structure-function relationships. Furthermore, SAXS results suggest conformational variation is a general functional feature of macromolecules. Thus, accurate structural analyses will require a comprehensive approach that assesses both flexibility, as seen by SAXS, and detail, as determined by X-ray crystallography and NMR. Here, we review recent SAXS computational tools, technologies, and applications to nucleic acids and related structures.

Solution structure of RNase P RNA

“…Fitting structure to solution envelope with EOM and NMA A number of computational techniques have emerged in recent years that allow improvement of atomic models to achieve a better agreement with observed SAXS data such as the ensemble optimization method (EOM) (Bernado et al 2007;Putnam et al 2007;Rambo and Tainer 2010a). In EOM, a large set of conformationally different models are generated from a set of starting models to generate a large pool of reasonable conformational alternatives; small ensembles of conformers are then selected from the pool by a Monte Carlo-based genetic algorithm to maximize the agreement between theoretical scattering by a selected ensemble and the experimental data.…”

Structure of the full-length HCV IRES in solution

“…Characterizing the structure of large flexible macromolecules remains, however, a challenge. Recent advances in small-angle scattering of X-rays (SAXS) or neutrons have turned these methods into powerful tools for characterizing the shape and accessible conformational space of flexible macromolecules and macromolecular assemblies 29,30 . Combining small-angle scattering data with other structural and biophysical experimental observations or with computational approaches led to the characterization of various proteins, RNAs and multi-molecular complexes 29 .…”

Protein grafting of p53TAD onto a leucine zipper scaffold generates a potent HDM dual inhibitor

“…4), supporting the notion that a high-affinity binding of p53LZ2 to HDMs is likely attributed to large binding areas through the N-terminal and C-terminal segments of p53LZ2 in part. As a third experiment, given the available crystal structures and possible protein complex models, we co-purified homodimeric p53LZ2 in complex with either HDM2NTD or HDMXNTD from E. coli and further investigated the resulting protein complexes using SAXS analysis [34][35][36] . All SAXS measurements of the p53LZ2-HDM complexes, in PBS buffer, were done in triplicate exposures and analysed to calculate the final composite scattering curves (Fig.…”