Regiospecific Flavonoid 7-O-Methylation with Streptomyces avermitilis O-Methyltransferase Expressed in Escherichia coli

Abstract

O-Methylation, commonly found in synthesis of secondary metabolites of plants and micro-organisms, appears to transfer a methyl group to the hydroxyl group of the recipient which increases the hydrophobicity of the recipient. O-Methyltransferase (OMT), , was isolated and characterized from Streptomyces avermitilis MA-4680. Its amino acid sequence showed 68% similarity with antibiotic C-1027 OMT and 53% similarity with the carminomycin 4-OMT. was expressed in E. coli as a His-tag fusion protein and showed that the methyl was transferred onto the 7-hydroxyl group of the isoflavones, daidzein and genistein, and the flavones, kaempferol and quercetin, as well as the flavanone naringenin. NMR and liquid chromatography-mass spectrometry were used to confirm the location of the methyl group on the recipient compound of naringenin, which was biotransformed into sakuranetin by E. coli transformant expressing (E. coli Sa-2). Therefore, E. coli Sa-2 would be used for the synthesis of the antifungal flavonoid, sakuranetin, through biotransformation.

Purification and Identification of Naringenin 7-O-Methyltransferase, a Key Enzyme in Biosynthesis of Flavonoid Phytoalexin Sakuranetin in Rice

“…In the case of F1-OMT in barley, the methylation activity on apigenin was three times higher than that on naringenin (20). In contrast, SaOMT-2 has broad substrate specificity and can catalyze the methylation of isoflavonoids as well as naringenin (21). Sakuranetin has not been identified in either barley or S. avermitilis.…”

“…Substrate Specificity of GST-OsNOMT-It was reported that F1-OMT and SaOMT-2 catalyze the methylation of several flavonoids, including naringenin (20,21). Therefore, we determined the substrate specificity of OsNOMT against several types of phenolic compounds, including flavonoids and isoflavonoids.…”

“…Although an O-methyltransferase (OMT) from barley (F1-OMT) that mainly methylates apigenin to form genkwanin has weak NOMT activity, sakuranetin has not been identified in barley (20). It was also reported that another OMT isolated from Streptomyces avermitilis (SaOMT-2) is able to catalyze the NOMT reaction, but SaOMT-2 has broad substrate specificity against isoflavones, flavones, and a flavanone, and its biological function remains unknown (21). Although sakuranetin is not found in healthy rice leaves, its biosynthesis is rapidly induced by both biotic and abiotic stresses, such as infection with phytopathogens such as M. oryzae, Xanthomonas oryzae, and Ditylenchus angustus, infestation with Sogatella furcifera, ultraviolet (UV) irradiation, and treatment with CuCl 2 or jasmonic acid (JA) (11,17,(22)(23)(24)(25).…”

Production of 7- O -Methyl Aromadendrin, a Medicinally Valuable Flavonoid, in Escherichia coli

“…The conversion of NRN to DHK was achieved in the presence of AT3G51240, but not with AT5G20400. On the other hand, it was reported that 7-O-methyltransferase from S. avermitilis (SaOMT) can efficiently convert isoflavanones, flavones, and flavanones, including NRN, into the corresponding 7-O-methylated derivatives (15). For this reason, we used the enzyme for further conversion of DHK into 7-OMA.…”

Plant Flavonoid O-Methyltransferases: Substrate Specificity and Application

“…Most of them are exclusive activity, where 7-OMTs that use flavonoids as a substrate do not use isoflavonoids as a substrate and vice versa. But SaOMT2 from Streptomyces avemilitis uses not only flavonoids but also isoflavonoids as a substrate with almost the same preference (Kim et al 2006a). It seems that OMTs from microorganisms contain more flexibility than those from plants in regards to substrate specificity (Kopycki et al 2008;Lee et al 2006).…”