sequencing data for qDMS-MaPseq
Functional RNA structures often contain long-range tertiary contacts that can determine whether an RNA forms its functional state. Understanding the thermodynamics of these tertiary contacts is crucial for decoding the folding and function of biological RNAs. Despite their importance, methods to measure tertiary contact thermodynamics are low-throughput or technically challenging, requiring specialized instruments. Here, we introduce a new quantitative dimethyl sulfate mutational profiling with sequencing (qDMS-MaPseq) method that leverages Mg2+ titrations to measure the thermodynamics of tertiary contacts in a high-throughput manner using reagents common to molecular biology labs. With qDMS-MaPseq, we measured the dG of unique 98 tetraloop/tetraloop receptor (TL/TLR) contacts with a single Mg2+ titration. These results agree well with measurements from specialized instruments (R2=0.64). DMS reactivity of individual nucleotides permitted observation of conformational changes in the TLR and the development of preliminary 3D models for some TLR mutants. These results indicate that qDMS-MaPseq can provide broad accessibility to tertiary contact thermodynamics at an unparalleled level of throughput.