Quorum Sensing-Based Detection and Biodegradation of Phthalate Esters by Engineered <i>Escherichia coli</i> and <i>Pseudomonas aeruginosa</i>

Phthalate esters (PAEs) are widely used in industry and have become prevalent toxic contaminants. Microbial biodegradation is promising, but its application is limited by the lack of efficient strains that can degrade a broad spectrum of PAEs. In this study, protein evolution was employed to generate a new variant of a PAE-degrading enzyme, <i>Go</i>Est15_V4, which exhibited 2.5- to 15-fold higher specific activity than the wild-type enzyme against <i>p</i>-nitrophenyl butyrate (pNPB) and nine other PAEs, including bulky substrates such as di(2-ethylhexyl) phthalate (DEHP), dihexyl phthalate, and diphenyl phthalate (DPHP). Additionally, autonomous whole-cell biodegraders containing a gene circuit XylS-QS-G_V4 (XylS(PA)-LuxI-LuxR-<i>Go</i>Est15_V4) were constructed, and the new engineered enzyme was expressed autonomously in <i>Escherichia coli</i> and <i>Pseudomonas aeruginosa</i> in response to PAEs through quorum sensing (QS). These strains degraded over 90% of diethyl phthalate and benzyl butyl phthalate in 12 h, and 80-90% of DEHP in 36-48 h, maintaining high efficiency over 5-round repeating cycles. The combination of protein engineering and QS enables the development of new whole-cell degrader strains with an enhanced degrading capacity and broad substrate tolerance.