peucetius and functionally characterized. Moreover, its biotechnological application for the production of diverse classes of O-methoxy natural products was experimented and investigated. Among these substrates both in the in vitro and in vivo bioconversion experiments, 7,8-dihydroxyflavone showed significant product conversion and was further analyzed by NMR study and confirmed as 7-hydroxy 8-O-methoxyflavone. Fermentation bioconversion assay was also performed with 7,8-DHF. Moreover, while analyzing the in vivo reaction mixtures by HR-QTOF-LC-ESI/MS, the mass fragments of O-methyl 3-hydroxyflavone, O-methyl phloretin, O-methyl luteolin, three mono- di- and tri-methylated genisteins, O-methyemodin and aloe-emodin were detected. Likewise, while analyzing the in vitro reaction mixture with sterol (β-sitosterol) and anthracyclines (doxorubicin and daunorubicin), the mass spectra of methoxy β-sitosterol, mono-, and di-O-methyl daunorubicin were obtained. This suggested the versatility of SpOMT7740 to accept different classes of natural products (flavonoids, sterols, anthracyclines, and anthraquinones) for the modification and production of O-methoxy compounds. These analyses indicate that OMT is non-regiospecific and capable of methylating the phenolic hydroxyl position of different classes of natural products as explained. Because it is an O-methyltransferase, the product detected with 3-hydroxyflavone should be 3-O-methoxyflavone and with sterol should be 3-O-methoxy β-sitosterol. Similarly, in the case of daunorubicin, the compound should be 6,11-di-O-methyldaunorubicin, but O-methyl daunorubicin could have attached at the 6- or 11- hydroxyl position. Moreover, the exact methylation position of O-methoxy emodin and aloe-emodin including O-methoxy genisteins, O-methoxy phloretin, and O-methoxy luteolin are
peucetius and functionally characterized. Moreover, its biotechnological application for the production of diverse classes of O-methoxy natural products was experimented and investigated. Among these substrates both in the in vitro and in vivo bioconversion experiments, 7,8-dihydroxyflavone showed significant product conversion and was further analyzed by NMR study and confirmed as 7-hydroxy 8-O-methoxyflavone. Fermentation bioconversion assay was also performed with 7,8-DHF. Moreover, while analyzing the in vivo reaction mixtures by HR-QTOF-LC-ESI/MS, the mass fragments of O-methyl 3-hydroxyflavone, O-methyl phloretin, O-methyl luteolin, three mono- di- and tri-methylated genisteins, O-methyemodin and aloe-emodin were detected. Likewise, while analyzing the in vitro reaction mixture with sterol (β-sitosterol) and anthracyclines (doxorubicin and daunorubicin), the mass spectra of methoxy β-sitosterol, mono-, and di-O-methyl daunorubicin were obtained. This suggested the versatility of SpOMT7740 to accept different classes of natural products (flavonoids, sterols, anthracyclines, and anthraquinones) for the modification and production of O-methoxy compounds. These analyses indicate that OMT is non-regiospecific and capable of methylating the phenolic hydroxyl position of different classes of natural products as explained. Because it is an O-methyltransferase, the product detected with 3-hydroxyflavone should be 3-O-methoxyflavone and with sterol should be 3-O-methoxy β-sitosterol. Similarly, in the case of daunorubicin, the compound should be 6,11-di-O-methyldaunorubicin, but O-methyl daunorubicin could have attached at the 6- or 11- hydroxyl position. Moreover, the exact methylation position of O-methoxy emodin and aloe-emodin including O-methoxy genisteins, O-methoxy phloretin, and O-methoxy luteolin are