Efficient hydroxylation of flavonoids by using whole-cell P450 sca-2 biocatalyst in Escherichia coli

Abstract

The hydroxylation is an important way to generate the functionalized derivatives of flavonoids. However, the efficient hydroxylation of flavonoids by bacterial P450 enzymes is rarely reported. Here, a bacterial P450 sca-2mut whole-cell biocatalyst with an outstanding 3′-hydroxylation activity for the efficient hydroxylation of a variety of flavonoids was first reported. The whole-cell activity of sca-2mut was enhanced using a novel combination of flavodoxin Fld and flavodoxin reductase Fpr from Escherichia coli. In addition, the double mutant of sca-2mut (R88A/S96A) exhibited an improved hydroxylation performance for flavonoids through the enzymatic engineering. Moreover, the whole-cell activity of sca-2mut (R88A/S96A) was further enhanced by the optimization of whole-cell biocatalytic conditions. Finally, eriodictyol, dihydroquercetin, luteolin, and 7,3′,4′-trihydroxyisoflavone, as examples of flavanone, flavanonol, flavone, and isoflavone, were produced by whole-cell biocatalysis using naringenin, dihydrokaempferol, apigenin, and daidzein as the substrates, with the conversion yield of 77%, 66%, 32%, and 75%, respectively. The strategy used in this study provided an effective method for the further hydroxylation of other high value-added compounds.

Rational Design of Key Enzymes to Efficiently Synthesize Phycocyanobilin in Escherichia coli

“…Twenty predicted atom models were scored, and the loop-region, main chain and side chain were refined; the HO T model with a −25997 of DOPE SCORE and 98.50% residues in the allowed region was selected as the best model. As a molecular docking method based on CHARMm’s position, CDOCKER can produce highly accurate docking results for various types of covalent and non-covalent interactions, and it has been successfully applied in the docking of small-molecule ligands to hemoproteins, such as cytochrome P450 [ 26 , 27 , 28 ]. The analysis of molecular docking in this study was performed using the CDOCKER tool in DS 2019.…”

Optimizing the Biosynthesis of Dihydroquercetin from Naringenin in Saccharomyces cerevisiae

“…17 By employing a novel combination of redox partners and enzyme engineering, along with optimized whole-cell biocatalytic conditions, a yield of 0.66 g/g DHQ was ultimately achieved in E. coli. 18 Furthermore, E. coli produced a remarkable yield of 239.4 mg/ L DHQ using glycerol as a carbon source, following optimization of the biosynthetic pathway and modification of the cytochrome P450 signal peptide. 19 In another study, the stability of F3′H in cells was improved by modifying predicted ubiquitination sites.…”

“…By employing a novel combination of redox partners and enzyme engineering, along with optimized whole-cell biocatalytic conditions, a yield of 0.66 g/g DHQ was ultimately achieved in E. coli . Furthermore, E.…”