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Evaluation of enzymatic degradation of petase against PBT and PEN nanoplastics using dye release assay
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Researchers expressed, purified, and tested wild-type and engineered variants of PETase enzyme against polyethylene naphthalate and polybutylene terephthalate nanoplastic substrates, developing a dye-release fluorescence assay that correlated with HPLC measurements to enable rapid, quantitative screening of enzymatic plastic degradation.
Plastic pollution poses a significant environmental challenge in recent years, driving interest in enzymatic degradation of plastic as one of the sustainable solutions. This study focuses on the expression, purification, and activity testing of different variants (WT, V3 and FAST) of PETase, an enzyme known to degrade polyethylene terephthalate (PET), using Escherichia coli (E. coli) SHuffle cells as the expression host. Following purification, the enzymatic activity of PETase was assessed on plastic-based nanoparticle substrates, specifically polyethylene naphthalate (PEN) and polybutylene terephthalate (PBT). Substrate degradation was monitored by a dye release assay using plastic nanoparticle- encapsulated dyes, Acridine Orange (AO) and Toluidine Blue O (TBO), providing an effective method for quantifying degradation. Fluorescence measurements obtained via spectrophotometry were found correlated with high-performance liquid chromatography (HPLC) data, which served as a reference standard for validation. This work contributes to the growing field of biocatalytic plastic degradation and highlights the potential for fluorescence-based assays in rapid enzyme screening.
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This study developed a simplified, high-yield preparation method for PET-degrading hydrolase enzymes to improve their ability to break down PET nano- and microplastics. The engineered enzyme showed enhanced hydrolysis activity against PET microplastics, offering a more practical route to enzymatic plastic waste treatment.
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Researchers developed a fast lab test that uses an enzyme to break down PET plastic particles into measurable pieces, detected through fluorescence. In water samples, the test measured both microplastics and much smaller nanoplastics, with results matching those from standard laboratory instruments.
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Researchers engineered PET-degrading enzymes to create variants that form covalent bonds with PET microplastic particles, enabling enzyme-based detection of microplastics. This proof-of-concept work repurposes plastic-degrading enzymes as analytical detection tools.
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