Fig. 4 in Genome-wide identification and characterization of genes involved in melatonin biosynthesis in Morus notabilis (wild mulberry)

Fig. 4. Purification and enzymatic analyses of recombinant His-tagged MnSNAT5. A: Purification of recombinant His-tagged MnSNAT5 (lane1: Protein Marker; lane2: pCold TF (+IPTG); lane3: recombinant (-IPTG); lane4: recombinant (+IPTG); lane5: purified protein). B: Mass spectrum of compounds detected using UPLCMS/MS. a: N-acetylserotonin standard; b: Chromatogram of serotonin catalyzed by MnSNAT5. C: Determination of Km and Vmax of MnSNAT5 for serotonin. MnSNAT5 (1.0 μg) was co-incubated with different concentrations of serotonin for 20 min at 37 ◦C. D: Mass spectrum of compounds detected using UPLC-MS/MS. a: standard, b: Chromatogram of 5-methoxytryptamine catalyzed by MnSNAT5. E: Determination of Km and Vmax of MnSNAT5 for 5-methoxytryptamine. MnSNAT5 (1.0 μg) was coincubated with different concentrations of 5-methoxytryptamine for 20 min at 37 ◦C. " IPTG" stand without addition of 1 mM IPTG to the E. coli BL21 (DE3) culture, "+IPTG" stand addition of 1 mM IPTG to the E. coli BL21 (DE3) culture. "TF" is Trigger Factor, a prokaryotic ribosome-related molecular chaperone that facilitates the translation and folding of peptides. "*" indicates Trigger factor. NAS: N-acetylserotonin; 5-MT: 5-methoxytryptamine; Mel: melatonin.