GENERAL STEPS: 30-50 μmol of 2-naphthol was dissolved in 0.5 mL of DMSO and diluted with buffer solution (50 mM Tris-HCl, pH 7.4, total volume 25 mL). Subsequently, 60-100 μmol of UDP-α-D-Glucose and 50 mL of AaGT3 crude enzyme solution extracted from wet-induced E. coli cells containing pET28a-AaGT3 were added. The reaction mixture was incubated at 30°C for no more than 12 hours. After completion of the reaction, five extractions were performed with 5 × 100 mL of ethyl acetate. The organic phases were combined and concentrated to dryness under reduced pressure. The residue was dissolved in 1.5 mL of methanol and purified by reversed-phase semi-preparative high-performance liquid chromatography (HPLC). The final product was structurally confirmed by mass spectrometry (MS) and nuclear magnetic resonance (NMR).
