FAD/Folate-Dependent tRNA Methyltransferase: Flavin as a New Methyl-Transfer Agent

RNAs contain structurally and functionally important modified nucleosides. Methylation, the most frequent RNA modification in all living organisms, mostly relies on SAM (S-adenosylmethionine)-dependent methyltransferases. TrmFO was recently discovered as a unique tRNA methyltransferase using instead methylenetetrahydrofolate and reduced flavin adenine dinucleotide (FAD) as essential cofactors, but its mechanism has remained elusive. Here, we report that TrmFO carries an active tRNA-methylating agent and characterize it as an original enzyme-methylene-FAD covalent adduct by mass spectrometry and a combination of spectroscopic and biochemical methods. Our data support a novel tRNA methylating mechanism.