RNA structures and dynamics with Å resolution revealed by x-ray free-electron lasers

Abstract

RNA macromolecules, like proteins, fold to assume shapes that are intimately connected to their broadly recognized biological functions; however, because of their high charge and dynamic nature, RNA structures are far more challenging to determine. We introduce an approach that exploits the high brilliance of x-ray free-electron laser sources to reveal the formation and ready identification of angstrom-scale features in structured and unstructured RNAs. Previously unrecognized structural signatures of RNA secondary and tertiary structures are identified through wide-angle solution scattering experiments. With millisecond time resolution, we observe an RNA fold from a dynamically varying single strand through a base-paired intermediate to assume a triple-helix conformation. While the backbone orchestrates the folding, the final structure is locked in by base stacking. This method may help to rapidly characterize and identify structural elements in nucleic acids in both equilibrium and time-resolved experiments.

All-Atom Biomolecular Simulation in the Exascale Era

“…The current explosive growth in computational resources coincides with a rapid development of advanced light sources, 52 neutron scattering facilities, 53 (in cell) NMR, 54 and cryoEM and cryo-electron tomography techniques, 55,56 all of which generate large data sets. For example, the LCLS-II free electron X-ray laser facility at SLAC has recently come online, with a nearly 10000-fold increase in pulse frequency compared to the existing LCLS beamline.…”

“…For example, the LCLS-II free electron X-ray laser facility at SLAC has recently come online, with a nearly 10000-fold increase in pulse frequency compared to the existing LCLS beamline. 52,57 Exascale computing will play a key role in guiding and analyzing the resulting high fidelity experiments that hold the potential to reveal biomolecular structure and dynamics, producing "movies" of complex molecular-level processes in action.…”

“…The current explosive growth in computational resources coincides with a rapid development of advanced light sources, neutron scattering facilities, (in cell) NMR, and cryoEM and cryo-electron tomography techniques, , all of which generate large data sets. For example, the LCLS-II free electron X-ray laser facility at SLAC has recently come online, with a nearly 10000-fold increase in pulse frequency compared to the existing LCLS beamline. , Exascale computing will play a key role in guiding and analyzing the resulting high fidelity experiments that hold the potential to reveal biomolecular structure and dynamics, producing “movies” of complex molecular-level processes in action.…”

Biophysical Investigation of RNA ⋅ DNA : DNA Triple Helix and RNA : DNA Heteroduplex Formation by the lncRNAs MEG3 and Fendrr

“…Furthermore, Zielinski et al . revealed the formation of RNA duplexes prior to the triplex formation by time‐resolved WAXS data, highlighting the dynamic nature of RNA structure [33] . To date, the DNA ⋅ DNA : DNA triplex formation has been demonstrated in vivo through the use of monoclonal antibodies labeled as “triplex‐specific”, including Jel318 and Jel466 [34,35] or with additional immunofluorescence staining, [35] as well as fluorescent molecules such as thiazole orange [36,37] .…”