On the specificity of the recognition of m6A-RNA by YTH reader domains

Most processes of life are the result of polyvalent interactions between macromolecules, often of heterogeneous types and sizes. Frequently, the times associated with these interactions are prohibitively long for interrogation using atomistic simulations. Here, we study the recognition of N6-methylated adenine (m$^{6}$A) in RNA by the reader domain YTHDC1, a prototypical, cognate pair that challenges simulations through its composition and required timescales. Simulations of RNA pentanucleotides in water reveal that the unbound state can impact (un)binding kinetics in a manner that is both model- and sequence-dependent. This is important because there are two contributions to the specificity of the recognition of the Gm$^{6}$AC motif: from the sequence adjacent to the central adenine and from its methylation. Next, we establish a reductionist model consisting of an RNA trinucleotide binding to the isolated reader domain in high salt. An adaptive sampling protocol allows us to quantitatively study the dissociation of this complex. Through joint analysis of a data set including both the cognate and control sequences (GAC, Am$^{6}$AA, and AAA), we derive that both contributions to specificity, sequence and methylation, are significant and in good agreement with experimental numbers. Analysis of the kinetics suggests that flexibility in both the RNA and the YTHDC1 recognition loop leads to many low-populated unbinding pathways. This multiple-pathway mechanism might be dominant for the binding of unstructured polymers, including RNA and peptides, to proteins when their association is driven by polyvalent, electrostatic interactions.