Fig. 1: Design and characterization of two BCHA biosensor configurations.

a Schematics of the biosensor in the isobutanol configuration (gray box) and simplified isobutanol pathway (enclosed by dashed lines; each arrow represents one enzymatic step). The α-isopropylmalate (α-IPM), which activates Leu3p (green dashed arrow), is an isobutanol by-product that accumulates due to deletion of LEU2. Thus, adding a PEST tag to destabilize yEGFP reduces background. α-KIV, α-ketoisovalerate. b Correlation between specific isobutanol titers (mg/L/OD) produced by engineered strains and GFP median fluorescence intensity (MFI) from the isobutanol biosensor. Strains used (from left to right): YZy121, YZy230, YZy81, YZy231, YZy232, YZy233, YZy234, YZy235, and YZy236 (Supplementary Table 1). c Schematics of the biosensor in the isopentanol configuration (gray box) and simplified isopentanol pathway (enclosed by dashed lines; each arrow represents one enzymatic step). In this case, α-IPM is an isopentanol precursor that does not accumulate due to LEU2 activity, so the PEST tag on yEGFP is not required. d Correlation between specific isopentanol titers (mg/L/OD) produced by engineered strains and GFP median fluorescence intensity (MFI) from the isopentanol biosensor. Strains used (from left to right): SHy187, SHy188, SHy192, SHy159, SHy176, and SHy158 (Supplementary Table 1). The MFI for each strain was measured 13 h after inoculation, during the exponential phase, and plotted against the corresponding specific isobutanol or isopentanol titers obtained after 48-h high-cell-density fermentations. MFI are represented in arbitrary units (arb. units). All data are shown as mean values. Error bars represent the standard deviation of at least three biological replicates. Source data are provided as a Source Data file.