Fig. 1: Engineering P. putida as a broadly applicable sensor for structurally diverse chemicals.

A General concept underlying whole-cell chemical detection by sensor strains. Rewiring the metabolism of P. putida enables selective assimilation of specific analytes (gray symbols), allowing for growth-dependent detection. The applications in this study (testing the efficiency of in vitro enzymatic activities, detecting industrially relevant metabolites produced by cell factories, and characterizing plastic breakdown by engineered bacteria in a co-culture system) are indicated in the diagram. B Metabolic rewiring of P. putida KT2440 to develop the first-generation SENS strain. Assimilation of glucose and its oxidized derivatives, gluconate and 2-ketogluconate (2KG), was blocked by eliminating genes encoding sugar transporters (gtsABCD, glucose transporter; gntT and PP_0652, gluconate transporters; and kguT, 2KG transporter), sugar kinases (glk, glucose kinase; gnuK, gluconate kinase; and kguK, 2KG kinase), and specific dehydrogenases (gcd, glucose dehydrogenase; and gad, gluconate dehydrogenase). Isocitrate lyase (aceA) and a key gene of the type VI secretion system (tssA1) were deleted in derivatives of the first-generation SENS strain for specific applications. The reporter module comprises a constitutively expressed msfGFP gene, inserted in the genome at the Tn7 attachment site (attTn7). Abbreviations: TCA cycle; tricarboxylic acid cycle; G6P, glucose-6-phosphate; 6PG, 6-phosphogluconate; 2K6PG, 2-keto-6-phosphogluconate; OAA, oxaloacetate; AKG, α-ketoglutarate; and CoA, coenzyme A. C Growth profile of wild-type (WT) P. putida KT2440 and the sensor SENS strain on different sugars. Both strains were incubated in de Bont minimal medium supplemented with either glucose, gluconate, or 2KG; cell density was estimated as the optical density at 600 nm (OD₆₀₀). Specific growth rates are indicated. D Fluorescence profile of strain SENS grown on different carbon sources supplemented at 10 mM. Cultures were prepared as indicated for panel (C); abbreviations: PCA, protocatechuate; AU, arbitrary units. In all cases, results represent average values ± standard deviations from three independent experiments.