Introducing Structure–Energy Concepts of RNA at the Undergraduate Level: Nearest Neighbor Thermodynamics and NMR Spectroscopy of a GAGA Tetraloop

Abstract

Two-dimensional nuclear magnetic resonance (2D NMR) techniques are regularly used for the determination of RNA structures. In an effort to introduce advanced level students who are generally not acquainted with or exposed to these techniques, this work, which has been designed as a practicum, utilizes a combination of 2D NMR techniques to help students gain necessary skills for RNA structure determination. The exercises presented here will help students fold RNA and predict the thermodynamic stable secondary structure using the independent nearest neighbor hydrogen bond (INN-HB) model and calculate the RNA folding free energies. This is usually the first step for structural determination of RNA. Students will then work with 1D NMR chemical shifts to confirm the base pairing pattern of the nucleotides in their secondary structure and finally analyze several 2D homo- and heteronuclear NMR data sets of a GAGA tetraloop. Overall, the practicum will give students a solid foundation for advanced RNA structure determination.