Fig. 5: Methyltransferase activity using a split aptamer of preQ1-I RNA.
From: A natural riboswitch scaffold with self-methylation activity
a Sequences of the two Tt preQ1-I fragments used and anion-exchange HPLC traces of the reaction mixture at time points 0 and 48 h (25 μM each RNA strand, 1.5 mM m6preQ1, 2.0 mM MgCl2, 100 mM KCl, MES buffer pH 6.0, 37 °C); as expected, the m3C-modified RNA product elutes earlier. b Secondary structure of the m3C-modified preQ1-stem-loop RNA and CAD-MS of co-isolated (M − 5H)5- ions of unreacted 21 nt RNA (61.1%, mexp. 6683.926 Da, mcalc. 6683.929 Da) and methylated RNA (38.9%, mexp. 6697.942, mcalc. 6697.945 Da) produced at 65 eV laboratory frame energy c and y fragments with and without methylation. c Thereby, the fraction of methylated c fragments (cm) increased from 0% to 36.8 ± 2.6% at site 15 (mean of values for sites 15 to 20 ± standard deviation). d The fraction of methylated y fragments (ym) decreased from 38.4 ± 1.4% to 0% at site 15 (mean of values for sites 1 to 14 ± standard deviation), which both are consistent with C15 as methylation site and with the yields independently obtained by HPLC analysis. Source data are provided in the Source Data file.
