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<p>Flavonoids are often decorated with methyl groups, which are catalyzed by flavonoid <i>O</i>-methyltransferases (FOMTs). Most FOMTs methylate flavonoids in a regiospecific manner. Because of the regiospecific nature of FOMTs, the synthesis of polymethoxyflavonoids is accomplished by multiple <i>O</i>-methylation steps. The multistep synthesis of dimethoxyflavonoids can be efficiently performed by a one-pot procedure using a multienzyme biocatalyst. For the one-pot production of dimethoxyflavonoids, fusion FOMTs were generated by the combination of two different regiospecific FOMTs. RdOMT10 (flavonoid 3-OMT), OsNOMT (flavonoid 7-OMT), and ObFOMT5 (flavonoid 4'-OMT) were used in the FOMT fusion. The fusion FOMTs (OsNOMT/ObFOMT5, OsNOMT/RdOMT10, and ObFOMT5/RdOMT10) were heterologously expressed in <i>Escherichia coli</i>. Activity assays of the recombinant fusion FOMTs demonstrated that OsNOMT/ObFOMT5, OsNOMT/RdOMT10, and ObFOMT5/RdOMT10 catalyze 7/4'-<i>O</i>-methylations, 7/3-<i>O</i>-methylations, and 4'/3-<i>O</i>-methylations of flavonoids, respectively. OsNOMT/ObFOMT5 and OsNOMT/RdOMT10 showed strong dimethylation activity towards diverse flavonoids and were therefore used in the site-selective bioconversion of flavonoids into dimethoxyflavonoids. The <i>E. coli</i> cells bearing OsNOMT/ObFOMT5 successfully converted flavonoids into 7,4'-dimethoxyflavonoids. The engineered <i>E. coli</i> expressing OsNOMT/RdOMT10 converted flavonoids into 3,7-dimethoxyflavonoids. This result indicates that the fusion FOMTs are useful multienzyme biocatalysts for the site-selective production of dimethoxyflavonoids by one-pot bioconversion.</p>

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Site-selective dimethylation of flavonoids using fusion flavonoid O-methyltransferases

  • 期刊论文

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  • 发表日期：

    2025年2月8日

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  • Kyungha Lee,
  • Seong Hee Bhoo,
  • Sang-Won Lee,
  • Man-Ho Cho

摘要

Flavonoids are often decorated with methyl groups, which are catalyzed by flavonoid O-methyltransferases (FOMTs). Most FOMTs methylate flavonoids in a regiospecific manner. Because of the regiospecific nature of FOMTs, the synthesis of polymethoxyflavonoids is accomplished by multiple O-methylation steps. The multistep synthesis of dimethoxyflavonoids can be efficiently performed by a one-pot procedure using a multienzyme biocatalyst. For the one-pot production of dimethoxyflavonoids, fusion FOMTs were generated by the combination of two different regiospecific FOMTs. RdOMT10 (flavonoid 3-OMT), OsNOMT (flavonoid 7-OMT), and ObFOMT5 (flavonoid 4'-OMT) were used in the FOMT fusion. The fusion FOMTs (OsNOMT/ObFOMT5, OsNOMT/RdOMT10, and ObFOMT5/RdOMT10) were heterologously expressed in Escherichia coli. Activity assays of the recombinant fusion FOMTs demonstrated that OsNOMT/ObFOMT5, OsNOMT/RdOMT10, and ObFOMT5/RdOMT10 catalyze 7/4'-O-methylations, 7/3-O-methylations, and 4'/3-O-methylations of flavonoids, respectively. OsNOMT/ObFOMT5 and OsNOMT/RdOMT10 showed strong dimethylation activity towards diverse flavonoids and were therefore used in the site-selective bioconversion of flavonoids into dimethoxyflavonoids. The E. coli cells bearing OsNOMT/ObFOMT5 successfully converted flavonoids into 7,4'-dimethoxyflavonoids. The engineered E. coli expressing OsNOMT/RdOMT10 converted flavonoids into 3,7-dimethoxyflavonoids. This result indicates that the fusion FOMTs are useful multienzyme biocatalysts for the site-selective production of dimethoxyflavonoids by one-pot bioconversion.