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Fluorescence‐activated screening of polyester‐depolymerizing enzymes based on pseudo‐PET polythioester plastics
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Researchers developed a novel fluorescence-based high-throughput screening strategy that detects sulfhydryl group release during enzymatic depolymerization of pseudo-PET polythioester plastics, enabling more direct identification of active polyester-degrading enzymes. The approach was successfully integrated with droplet microfluidics and validated using leaf-branch compost cutinase, offering a more efficient pipeline for discovering plastic-degrading enzymes and microorganisms.
Abstract While fluorescence‐based high‐throughput screening (HTS) has been valuable for identifying enzymes and microorganisms for polyethylene terephthalate (PET) depolymerization, the traditional approach relies on cleaving ester bonds in small‐molecule probes, limiting its ability to directly detect activity towards polymeric PET substrates. This study proposes a novel fluorescence‐based screening strategy for detecting the release of sulfhydryl groups during the depolymerization of pseudo‐PET polythioesters by polyester hydrolases. The strategy successfully identifies the polyester‐depolymerizing activity of leaf‐branch compost cutinase (LCC ICCG ), whereas porcine liver esterase (PLE) only hydrolyses small molecular substrates. Furthermore, the strategy can be integrated with a droplet microfluidic system for high‐throughput screening of LCC ICCG . This work further demonstrates the applicability of the microplate reader platform for screening polyester‐depolymerizing bacteria. This novel fluorescence‐based approach offers an efficient strategy to identify enzymes and microbial resources for depolymerizing polyester‐like plastics.
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