Nonadditive Conformational Effects of tRNA Core Modifications

Post-transcriptional modifications are known to regulate the structure and function of tRNA molecules. Compared to modifications in the anticodon region, the mechanistic effects of modifications in the core region of tRNA remain poorly understood. Using all-atom molecular dynamics (MD) simulations on yeast tRNAPhe, we investigated the conformational effects of incorporating chemical modifications in the core region (termed hereafter "core modifications") of tRNA. Specifically, we focused on three core modifications, namely, 7-methylguanosine (m7G), 5-methylcytosine (m5C), and 1-methyladenosine (m1A). We discovered that individual modifications increased the structural stability of tRNA, but their combinations produced multimodal free energy landscapes with multiple distinct basins. We identified m1A58 as a bifunctional regulator that locally disrupts structure while globally coordinating anticorrelated arm movement. The combination of m1A58 and m5C49 exhibited the lowest structural deviation from the crystal structure, while the triply modified variant incorporating all three modifications produced the most integrated residue community network, reflecting nonadditive allosteric coupling that cannot be predicted from any pairwise combination. Critically, core modifications established extensive allosteric pathways that prevent the consistent inward displacement of the anticodon stem loop otherwise observed in all simulations of chemically unmodified tRNA. Our results demonstrate that core modifications function as regulators of conformational ensembles rather than simple stabilizers.