Extended Data Fig. 2: Time-course studies of the SoBcmB catalytic reactions using 3 and 4 as substrates.

a and e, Proposed steps of SoBcmB-catalysed reactions using 3 and 4 as substrates, respectively. b and f, HPLC analyses of the SoBcmB catalytic reactions. The enzymatic reaction (50 μl) containing 50 mM Tris–HCl buffer (pH 7.5), 0.6 mM substrate, 2 mM αKG, 2 mM l-ascorbic acid, 50 μM FeSO₄·7H₂O, and 50 μg purified enzyme (27 μM) was incubated at 37 °C. The reactions were quenched at different times by the addition of 100 μl of precooled methanol and centrifuged at 14,000g for 20 min. The supernatants were analysed by HPLC and LC/MS (Thermo Fisher LTQ Fleet mass spectrometer). Three experiments were repeated independently with similar results. c and g, UV-Vis spectra for 3 and 3a–c and 4, 4a and 4c. d and i, ( + )-ESI-MS spectra of 3, 3a–c, 4, and 4a–4c. h, Extracted ion chromatogram (EIC) corresponding to intermediate 4b in SoBcmB-catalysed reactions. The LC/MS analysis was conducted by Thermo Scientific Q Exactive Plus Orbitrap LC–MS/MS System.