Fig. 4. Design of a hydroxytyrosol biosensor and its application in TYO activity optimization.

a Design of a hydroxytyrosol biosensor and the strategy of applying it in in vivo-directed evolution of TYO for improved hydroxytyrosol biosynthesis. b β-Galactosidase activity (blue) and hydroxytyrosol productions (green) in strain H2 harboring control plasmid and plasmid P6, after culturing at 37 °C for 36 h. c Hydroxytyrosol productions in the wild-type and selected clones in the first and second rounds of in vivo-directed evolution, after culturing at 37 °C for 36 h, in the presence of 1 mM tyrosine; Time courses of productions of hydroxytyrosol and dopamine, as well as tyrosine consumption, from strain BHYT carrying plasmid P6 (d), P8 (e), or P9 (f) in a 5-L fermentor. Blue circle, brown triangle and green triangle indicate the concentration of tyrosine, dopamine and hydroxytyrosol, respectively. g Specific activities of wild-type and mutant TYOs. The data shown in b, c, d, e, f and g are from three replicate experiments and are expressed as the mean±SD. Source data underlying Fig. 4b-g are provided as a Source Data file.