tRNA molecules get heavily modified post-transcriptionally. The N-1 methylation of purines at position 9 of eukaryal and archaeal tRNA is catalyzed by the SPOUT methyltranferase Trm10. Remarkably, while certain Trm10 orthologs are specific for either guanosine or adenosine, others show a dual specificity. Structural and functional studies have been performed on guanosine- and adenosine-specific enzymes. Here we report the structure and biochemical analysis of the dual-specificity enzyme from (Trm10). We report the first crystal structure of a construct of this enzyme, consisting of the N-terminal domain and the catalytic SPOUT domain. Moreover, crystal structures of the SPOUT domain, either in the apo form or bound to -adenosyl-l-methionine or -adenosyl-l-homocysteine reveal the conformational plasticity of two active site loops upon substrate binding. Kinetic analysis shows that Trm10 has a high affinity for its tRNA substrates, while the enzyme on its own has a very low methyltransferase activity. Mutation of either of two active site aspartate residues (Asp206 and Asp245) to Asn or Ala results in only modest effects on the N-1 methylation reaction, with a small shift toward a preference for mG formation over mA formation. Only a double D206A/D245A mutation severely impairs activity. These results are in line with the recent finding that the single active-site aspartate was dispensable for activity in the guanosine-specific Trm10 from yeast, and suggest that also dual-specificity Trm10 orthologs use a noncanonical tRNA methyltransferase mechanism without residues acting as general base catalysts.