Covalent modifications of standard DNA/RNA nucleobases affect epigenetic regulation of gene expression by modulating interactions between nucleic acids and protein readers. We derive here the absolute binding free energies and analyze the binding modalities between key modified nucleobases 5-methylcytosine (5mC), 5-hydroxymethylcytosine (5hmC) and N6-methyladenine (m6A) and all non-prolyl/non-glycyl protein side chains using molecular dynamics simulations and umbrella sampling in both water and methanol, the latter mimicking the low dielectric environment at the dehydrated nucleic-acid/protein interfaces. We verify the derived affinities by comparing against a comprehensive set of high-resolution structures of nucleic-protein complexes involving 5mC. Our analysis identifies protein side chains that are highly tuned for detecting cytosine methylation as a function of the environment and can thus serve as microscopic readers of epigenetic marks. Conversely, we show that the relative ordering of sidechain affinities for 5hmC and m6A does not differ significantly from those for their precursor bases, cytosine and adenine, respectively, especially in the low dielectric environment. For those two modified bases, the effect is more nuanced and manifests itself primarily at the level of absolute changes in the binding free energy. Our results contribute towards establishing a quantitative foundation for understanding, predicting and modulating the interactions between modified nucleic acids and proteins at the atomistic level.
© 2022. The Author(s).