In this study, we studied the biochemical characterization of flavone synthase I from Daucus carota (DcFNS I) and applied it with flavonoid 6-hydroxylase from Scutellaria baicalensis (SbCYP) to convert flavanones to flavones. The recombinant DcFNS I was expressed in the form of the glutathione-S-transferase fusion protein. Rather than taxifolin, naringenin, pinocembrin, and eriodictyol were accepted as substrates. The optimal temperature and pH for reaction in vitro were 35 °C and 7.5, respectively, and 2-oxoglutarate was essential in the assay system. Co²⁺, Cu²⁺, Mn²⁺, Ni²⁺, and Zn²⁺ were not substitutes for Fe²⁺. EDTA and pyruvic acid inhibited the activity, except for Fe³⁺. Kinetic analysis revealed that the V_max and k_cat values of the recombinant DcFNS I against naringenin were 0.183 nmol mg^-1 s^-1 and 0.0121 s^-1, and 0.175 nmol mg^-1 s^-1 and 0.0116 s^-1 against pinocembrin. However, the recombinant DcFNS I had a higher affinity for naringenin than pinocembrin, with k_M values for each of 0.076 mM and 0.174 mM respectively. Thus, it catalyzed naringenin more efficiently than pinocembrin. Subsequently, using an Escherichia coli and Saccharomyces cerevisiae co-culture system, we successfully converted naringenin and pinocembrin to scutellarein and baicalein respectively. In a synthetic complete medium, the titers of scutellarein and baicalein reached 5.63 mg/L and 0.78 mg/L from 200 mg/L precursors.

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Journal Article

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Zhang X, Qi Z, Fan X, Zhang H, Pei J, Zhao L

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