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. 2023 Feb 16;51(5):2434–2446. doi: 10.1093/nar/gkad081

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© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com

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Figure 2. — Key residues of CeMETT10- MTD for RNA methylation. (A) The structure of human METTL16-MTD (hMETTL16-MTD) in complex with hMAT2A- hp1 RNA (PDB: 6DU4) (23). hMAT2A-hp1 is shown in green. (B) RNA docking onto the CeMETT10-MTD structure. The structure of hMETTL16-MTD complexed with hMAT2A-hp1 in (A) was superimposed on the structure of CeMETT10-MTD. hMETTL16-MTD was omitted for clarity. hMAT2A-hp1 is colored green. (C) Methylation of sams-hp RNA (see Figure 3B) by METT10-MTD and its variants. The sams-hp RNA (1 μM) was incubated with 0.4 μM METT10-MTD in the presence of 1 mM SAM, at 37°C for 4 min. The methylation of sams-hp RNA by wild-type METT10-MTD was taken as 1.0. The bars in the graphs are the SD of three independent experiments. (D) Model of the hMETTL16 active site with SAH and hMAT2A-hp1. The SAH was modeled into the structure of hMETTL16-MTD in complex with hMAT2A-hp1 by referring to the structure of hMETTL16-MTD in complex with SAH (PDB: 6B92) (34). The methylation site in the RNA (m⁶A site) is shown in green, and SAH is colored magenta. (E) Superimposition of the active site structures of CeMETT10 (cyan) and hMETTL16 (orange). (F) Methylation of sams-hp RNA by METT10-MTD and its variants, as in (C). The methylation of sams-hp RNA by wild-type METT10-MTD was taken as 1.0. The bars in the graphs are the SD of three independent experiments. (G) Superimposition of the K-loops of CeMETT10-MTD (cyan) and hMETTL16-MTD (orange) in complex with hMAT2A-hp1. SAH was modeled as in (D). (H) Methylation reactions of sams-hp-ls RNA by METT10-MTD and its variants with various concentrations of SAM (0.05, 0.1, 0.2, 0.5, 1.0 and 1.5 mM). The sams-hp RNA (20 μM) was incubated with 0.4 μM METT10-FLΔL or its variants in the presence of various concentrations of SAM, at 37°C for 4 min. The bars in the graphs are the SD of three independent experiments.