Hippurate

For each circuit separate colonies of E. coli TOP10 (strain K-12, genotype: F⁻mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 recA1 araD139 Δ(ara-leu)7697 galU galK rpsL (Str^R) endA1 nupG) strains harboring the circuit plasmids were cultured overnight at 37 °C in LB with appropriate antibiotic. The next day each culture was diluted 100× in LB with antibiotics. 95 µL of fresh cultures were distributed in 96-well plate (Corning 3603) and the plate was incubated to reach the OD₆₀₀ ~ 0.1 in a plate reader (Biotek Synergy HTX). Then 5 µL of the input metabolites (100× ethanol solutions 5x diluted in LB) were added and the plate was incubated for 18 h at 37 °C. During the incubation, the OD₆₀₀ and GFP fluorescence (gain: 35, ex: 458 nm, em: 528 nm) were measured. Benzoate, hippurate, cocaine hydrochloride, benzaldehyde, benzamide and biphenyl-2,3-diol (2,3-dihydroxy-biphenyl) were purchased from Sigma-Aldrich. Permission to purchase cocaine hydrochloride was given by the French drug regulatory agency (Agence Nationale de Sécurité du Médicament et des Produits de Santé). For all chemicals, serial dilutions of 100× concentrations were prepared in ethanol. The formula presenting the results of the circuits’ characterization is shown in data normalization section. The mean and standard deviation of all normalized data are provided in Supplementary Table 7.