Fig. 3.
Engineering Gal3p to respond to xylose. a Design of the screen and selection systems. Gal3p expressed under its native promoter, in the presence of activation sugar (galactose/xylose), would activate the GAL regulon and switch ON GAL1p and GAL10p to drive expression of KANMX and EGFP, resulting in G418 sulfate resistance (selection) and fluorescence (screen). b Gal3p–WT interaction with galactose and xylose measured using the fluorescence assay. VEG16 transformed with selection and screening construct was grown in sucrose with varying concentrations of galactose (2% – 2 × 10−6%) or xylose (8% – 2 × 10−3%). c Fold-change in fluorescence of the best Gal3p mutants when incubated with varying concentrations of xylose. d Fold-change in fluorescence of the best Gal3p mutants when incubated with varying concentrations of galactose. Each data point represents average of three individual replicates ± sd