Extended Data Fig. 8: A comparison of the substrate-binding sites in two structurally characterized RS methylases.

a, b, The substrate complexes of TokK (a) and RlmN (PDB ID: 5HR7) (b) are shown. RlmN is an RS methylase that uses a radical-based mechanism to methylate an sp²-hybridized carbon (C2) of an adenine base in transfer or ribosomal RNA. By contrast to TokK and other Cbl-dependent RS enzymes, RlmN uses a 5′-dA• to activate a post-translationally modified methyl-Cys residue on a C-terminal loop in the active site to modify its aromatic substrate via radical addition. Comparison of a structure of RlmN in a cross-linked methylCys-tRNA intermediate state to the TokK substrate complex shows that, despite the differences in reaction mechanism, these systems use a similar orientation of radical initiator (5′-dA•), substrate target carbon (C6, C2), and methyl donor (OH, Me). In the TokK substrate complex, the -OH ligand of the Cbl cofactor serves as a surrogate for the position of the methyl donor.