Fig. 4: Engineering TPA assimilation in P. putida SENS.

A Polyethylene terephthalate (PET) can be degraded by hydrolytic enzymes (for instance, PETases and cutinases). Metabolites resulting from these enzymatic reactions are BHET, bis(2-hydroxyethyl) terephthalate; MHET, mono(2-hydroxyethyl) terephthalate; TPA, terephthalate; and EG, ethylene glycol. MHET can be converted into TPA and EG by MHETase; this enzyme also displays residual activity on BHET, yielding MHET and EG. Abbreviations: CoA, coenzyme A; DCD, 1,6-dihydroxycyclohexa-2,4-diene-dicarboxylate. B The tph operon from P. umsongensis, encoded in plasmid pBT·T_tph, enables TPA transport and catabolism in P. putida SENS·T. The tph operon encodes an IclR-type regulator (regulating the expression of the remaining genes in the cluster), the TPA 1,2-dioxygenase subunits α (tphA2) and β (tphA3), a reductase (tphA1), a dehydrogenase (tphB), and a TPA transporter (tphK). These activities convert TPA into protocatechuate (PCA). PCA is assimilated by P. putida through its conversion into succinate and acetyl-CoA. C Assessing TPA assimilation by the sensor P. putida SENS·T strain. P. putida SENS was transformed with either the pBT·T_tph plasmid or an empty vector (pSEVA221). The resulting strains were incubated in DBM medium supplemented with 2 g L^–1 TPA as the only carbon substrate. Growth and msfGFP fluorescence profiles are shown as average values ± standard deviation of three independent experiments. Abbreviations: AU, arbitrary units.