Fig. 7: A single amino acid substitution holds the key to dimethylation ability.

a Superposition of PsiM (beige with key residues Asn183 and Asn247 in green) and the METTL16-SAH complex (chain A of PDB 6GFK, grey). Residue numbers are shown in normal print and italics, respectively. The methyl group in baeocystin forces Asn183 of PsiM to shift to the left (arrow). Asn247 provides the necessary space and in addition stabilises the alternative conformation of Asn183 through a hydrogen bond (dashed line). A comparable movement of the corresponding residue in METTL16 (Asn184) is prevented by the presence of Met248 (double arrow) in the otherwise strictly conserved protein core. b Alignment of the region around N247 in PsiM with the sequences of other METTL16- and RlmF-like transferases. c Experimentally determined (di)methylation activity of wild-type PsiM and the N247M mutant. Shown are single ion chromatograms of norbaeocystin (red chromatogram, m/z 257 [M + H]⁺), baeocystin (grey, m/z 271 [M + H]⁺) and psilocybin (blue, m/z 285 [M + H]⁺). Top lane: overlaid chromatograms of authentic standards. Second lane: negative control with heat-denatured enzyme.