A practical guide to RNA NMR spectral analysis for undergraduate students

Nuclear magnetic resonance (NMR) spectroscopy is a powerful analytical tool that can be used to determine the structures of both small organic molecules and large biomacromolecules. Many undergraduate students are exposed to NMR spectral analysis during organic chemistry courses, but few students have the opportunity to learn about biomacromolecular NMR spectral analysis. Here, we present a three-part practicum, aimed at providing resources for faculty to introduce RNA structure and NMR analysis into existing upper-level courses. Students will begin by predicting an RNA secondary structure using a webserver that uses nearest neighbor parameters to estimate the folding stability. Then, students can verify the secondary structure using 1D NMR data. Students can then learn the iterative process of chemical shift assignments, through the concurrent analysis of multiple 2D NMR spectra. Students will also be able to visualize the RNA tertiary structure, allowing them to connect the somewhat abstract NMR spectra with measurable interproton distances. Collectively, the activities outlined in this practicum will help students develop representational competence and visual literacy skills and will ensure that they have a strong foundation for reading and evaluating data presented in the primary scientific literature.