Fluorescence-based high-throughput screening approaches facilitate the discovery of enzymes and microorganisms for polyethylene terephthalate (PET) depolymerization and recycling. However, the traditional method of activating the fluorescence signal by cleaving the ester bond on small molecule probes has limited ability in detecting enzymatic activity towards polymeric substrates. This study proposes a novel fluorescence-based screening strategy that detects the release of sulfhydryl groups during the depolymerization of pseudo-PET polythioesters by polyesterases. The strategy successfully identifies the polyester-depolymerizing activity of leaf-branch compost cutinase (LCC ICCG), while porcine liver esterase (PLE) only hydrolyses small molecular substrates. When combined with a droplet microfluidic system, the strategy enables high-throughput screening of LCC ICCG. The study also demonstrates that screening for polyester-depolymerizing bacteria can be performed via a microplate reader platform. The new screening approach offers an efficient method for identifying enzymes and microbial resources for depolymerizing polyester-like plastics.

Constructing multi-dimensional hydrogen bond (H-bond) regulated single-molecule systems with multi-emission remains a challenge. Herein, we report an excited-excited intramolecular transfer (ESIPT) featured chromophore (HBT-DPI) that shows flexible emission tunability via the multi-dimensional regulation of intra- and intermolecular H-bonds. The feature of switchable intramolecular H-bonds is induced via incorporating two hydrogen bond acceptors into a single-molecule system, HBT-DPI, allowing the “turn on/off” of ESIPT process by forming isomers with distinct intramolecular H-bonds. In response to different solvent environments, the obtained four types of crystal/cocrystals vary in the contents of isomers and the molecular packing modes, which are mainly guided by the intermolecular H-bonds, exhibiting non-emissive features or emissions ranging from green to orange. Moreover, we demonstrate the practical utility of this fluorescent material for visualizing hydrophobic/hydrophilic areas on large-scale heterogeneous surfaces of modified PVDF membranes and quantitatively estimate the surface hydrophobicity, providing a new approach for hydrophobicity/hydrophilicity monitoring and measurement.

A rapid and convenient strategy to monitor the productivity of biomanufacturing is essential for the research in optimizing relevant bioprocesses. In this work, we have developed a fluorescein-derived probe (FL-DT) that reacts rapidly with thiol groups via 1, 4-Micheal addition reaction of the sulfhydryl to unsaturated ketone and releases fluorescence. FL-DT specifically forms fluorescent adduct with two adjacent thiols in a protein of interest (POI), making the probe a reliable tool for protein quantification. The production of xylanase fused with a short di-Cys tag was then successfully monitored and quantified with FL-DT in E. coli system under different protein expression conditions, providing useful information for optimizing the bioprocess. Our work provides a convenient and efficient strategy for POI labeling and monitoring bioproduction.