Abstract
NMR structure calculation is inherently integrative, and can incorporate new experimental data as restraints. As RNAs have lower proton densities and are more conformational heterogenous than proteins, the refinement of RNA structures can benefit from additional types of restraints. Paramagnetic relaxation enhancement (PRE) provides distance information between a paramagnetic probe and protein or RNA nuclei. However, covalent conjugation of a paramagnetic probe is difficult for RNAs, thus limiting the use of PRE NMR for RNA structure characterization. Here, we show that the solvent PRE can be accurately measured for RNA labile imino protons, simply with the addition of an inert paramagnetic cosolute. Demonstrated on three RNAs that have increasingly complex topologies, we show that the incorporation of the solvent PRE restraints can significantly improve the precision and accuracy of RNA structures. Importantly, the solvent PRE data can be collected for RNAs without isotope enrichment. Thus, the solvent PRE method can work integratively with other biophysical techniques for better characterization of RNA structures.
Highlights
RNA is an important class of biological macromolecules
We show that the integrative use of NMR sPRE restraints greatly improves the precision and accuracy of RNA structures
Unlike the more established Paramagnetic relaxation enhancement (PRE) measurement, the sPRE is measured without the covalent conjugation of a paramagnetic probe
Summary
RNA is an important class of biological macromolecules. Different from protein-coding messenger RNAs, noncoding RNAs play important structural, signaling, and catalytic roles, and are involved in all aspects of cellular. Zhou Gong and Shuai Yang contributed to this work. The three-dimensional (3D) structures of non-coding RNAs are important for their functions (Butcher and Pyle 2011; Larsen et al 2019). The RNA folding is basically hierarchical (Herschlag et al 2018). Though there are programs to predict RNA secondary structures regarding the patterns of base-pairing (Bellaousov et al 2013; Parisien and Major 2008; Zhang et al 2019; Zuker 2003), it remains difficult to predict RNA secondary structures with complex topologies (Schlick and Pyle 2017; Zhao et al 2018)
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