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Updated: Jun 24, 2026

Probing RNA Structure with Dimethyl Sulfate Mutational Profiling with Sequencing In Vitro and in Cells
Published on: December 9, 2022
Probing mutation-induced structural perturbations by refinement against residual dipolar couplings: application to
John P Kirkpatrick1, Ping Li, Teresa Carlomagno
1Structural and Computational Biology, European Molecular Biology Laboratory, Heidelberg, Germany.
Abstract:
Confident interpretation of biochemical experiments performed with mutated proteins relies on verification of the integrity of the mutant structures. We present a simple and rapid refinement protocol for comparing the structures of mutated and wild-type proteins. Our approach involves measurement of residual dipolar couplings, and only requires assignment of the backbone resonances of the mutant species. We demonstrate application of the protocol to a mutant of the 15.5K protein, a core component of the U4 spliceosomal ribonucleoprotein (RNP) complex. Confirmation of the unperturbed structure of the mutated protein prompted re-examination of a previous mutagenesis study and indicated that the interpretation of mutant binding affinities in terms of direct interfacial contacts should be applied with caution.
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