Plastic pollution has emerged as a critical global environmental issue, with polyethylene terephthalate (PET) representing one of the most persistent and widely used plastics. Current enzymatic degradation methods for PET exhibit limited efficiency, hindering their application on an industrial scale(Guo etal,2025; V etal, 2020). To address this challenge, we aim to develop enhanced enzymatic strategies for PET biodegradation and recycling by screening and engineering high-efficiency plastic-degrading enzymes. Our objective is to improve the catalytic conversion of PET into its monomers, terephthalic acid (TPA) and ethylene glycol (EG), thereby providing a sustainable and innovative bioremediation solution(Fig 1).

Figure 1. Technical Roadmap

Design and Optimization of Part Cycle:

Cycle 1: PETase derived from Ideonella sakaiensis (positive control), capable of converting PET into MHET.

Cycle 2: Screening mutants of PET-degrading enzyme derived from deep-sea sources, capable of converting PET into MHET.

Cycle 3: MHETase derived from Ideonella sakaiensis, which converts MHET into TPA and EG. The mutant PETase screened in Cycle 2 is used in combination with the MHETase to degrade PET into recyclable TPA and ethylene glycol (EG).

The resulting engineered enzymes are expected to demonstrate significantly improved PET degradation performance, with potential applications in environmental remediation and industrial plastic waste processing.

Design:

A bacterial strain Ideonella sakaiensis capable of efficiently degrading PET has been discovered. At the same time, a hydrolase secreted by this bacterium with the ability to degrade PET, named PETase, has also been reported. This enzyme can hydrolyze PET into bis-2-hydroxyethyl-terephthalate (BHET), mono-2-hydroxyethyl-terephthalate (MHET), and TPA (YOSHIDA etal,2016).The genes encoding IsPETase fused with 6×His-tag at its C-terminus were optimized and chemically synthesized. The plasmid map was constructed by snapgene software(Fig. 2), and inserted into the expression vector pET28a by homologous recombination method, the recombinant plasmids pET28a-IsPETase were further transformed into E.Coli DH5α.

Fig 2. The plasmid map of pET28a-IsPETase

Build:

The pET-28a plasmid vector was linearized for the construction of the expression plasmid. The plasmid containing the IsPETase gene was constructed using homologous recombination, followed by confirmation of successful construction via agarose gel electrophoresis. The Fig 3A showed that the IsPETase gene is between 500bp and 1000bp markers. The results are showed in Fig 3B, the linearized plasmid pET28a is highlighted 5000bp. The expansion length of gene is consistent with the DNA encoding and indicate the succeed of amplification. The Fig 3C showed that the recombinant plasmids was transformed into processed E.coli DH5α to copy the plasmids. As shown in Fig 3D, the sequencing results revealed a perfect match to the target gene sequence without any mutations, demonstrating the successful construction of the plasmid pET28a-IsPETase.

Fig. 3.The electrophoresis, plate Assay, and sequencing of IsPETase. Note: IsPETase：879bp; pET28a：5323bp

Test：

We measured the OD600 at various time points. As shown in Figure 4A, no significant difference was observed between the growth curves of the control group and the IsPETase-expressing strain, indicating robust bacterial growth.

We engineered the expression construct to incorporate a specific gene sequence encoding a hexahistidine (His-tag) affinity tag, enabling ribosomal synthesis of the recombinant protein with a C-terminal His-tag. The His-tagged protein was subsequently purified using Ni-NTA affinity chromatography, whereby the immobilized nickel ions selectively bind the His-tag. Non-specifically bound proteins were removed by washing with buffer, and the target protein was eluted under native conditions using imidazole-containing elution buffer.

In SDS-PAGE analysis, the crude protein dsPETase exhibited diffuse banding patterns, indicative of the presence of non-target cellular proteins. Following purification, the Fig 4B result demonstrated that the molecular weight of the recombinant protein was between 25 kDa and 35 kDa, which is consistent with its theoretical molecular mass of 33.5 kDa.

Fig 4.The growth curve and SDS-PAGE of IsPETase

Learn:

In cycle 1, the plasmids are constructed successfully and transformed in E.coli. The SDS polyacrylamide gel electrophoresis (SDS-PAGE)experiment we conducted yields the expected proteins’ sizes of IsPETase, which indicates that the proteins IsPETase are successfully expressed in E.coli. This cycle experiments and proves the positive control, providing essential results for further designs.

Gly8-GIn(BBa_2542VYML), Gly8-Ser(BBa_25N2AZ4A), Gly8-Thr(BBa_25655FKG), Gly8-Cys(BBa_25EL7FQM), Gly8-Asn(BBa_25NSRU73), Gly8-Tyr(BBa_2578647W), Arg47-Lys(BBa_25VTOT4S), and Arg47-His(BBa_2543N0QT)

Design:

A novel halophilic PET plastic-degrading enzyme with high activity derived from the hydrothermal vent metagenome (of deep-sea origin) was reported. Meanwhile, a hydrolase with PET-degrading activity, namely PETase, secreted by this bacterium was also reported. This enzyme can hydrolyze PET into bis(2-hydroxyethyl) terephthalate (BHET), mono(2-hydroxyethyl) terephthalate (MHET), and terephthalic acid (TPA) [KANELLI et al,2025].

The genes encoding dsPETase fused with 6×His-tag at its C-terminus were optimized and chemically synthesized. The plasmid map was constructed by snapgene software(Fig 5), and inserted into the expression vector pET28a by homologous recombination method, the recombinant plasmids pET28a-dsPETase were further transformed into E.Coli DH5α.

Fig 5. The plasmid map of pET28a-dsPETase

To identify key active sites in dsPETase for mutagenesis, we constructed a three-dimensional protein structure of dsPETase and performed molecular docking with the chemical structure of PET(Fig 6). The docking score (binding energy) serves as an indicator to evaluate the affinity between PET and protein dsPETase. A stronger affinity results in tighter binding and more stable molecular interactions. A more negative docking score corresponds to a stronger affinity between the molecules.

Fig 6. Docking results of dsPETase with the PET molecule; Red represents the PET molecule.

Fig 7. Schematic diagram of the active site of dsPETase with the PET molecule.

The optimal binding mode between the PET molecule and dsPETase is shown in Figs 6 and 7. The molecular docking binding energy was calculated to be −3.249 kcal/mol, with stabilization primarily achieved through hydrogen bonding (Table 1). Key amino acid residues forming hydrogen bonds with the PET molecule were glycine at position 8 (Gly8) and arginine at position 47 (Arg47), which enhanced the stability of the PET molecule within the binding pocket.The molecular docking results validated the feasibility of selecting Gly8 and Arg47 for mutagenesis, a decision initially made based on expert opinions during the experimental design phase. However, experimental validation is required to confirm the degradation efficiency of the mutants and ensure the reliability of the result.

Based on the advice of biology Professor Yao, we performed site-directed mutagenesis on the active site residues glycine (Gly8) and arginine (Arg47). Specifically, Gly8 was mutated to amino acids with similar properties—namely, polar neutral R-group residues such as glutamine (Gln), serine (Ser), threonine (Thr), cysteine (Cys), asparagine (Asn), and tyrosine (Tyr). This strategy helps minimize structural perturbations that could lead to loss of protein activity. The arginine at position 47 was mutated to amino acids with similar properties, specifically basic R-group residues such as lysine (Lys) and histidine (His). The resulting mutants were designated as Arg47-Lys, Arg47-His, Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr. As shown in Table 1, the docking scores of these mutants were significantly improved compared to that of wild-type dsPETase, indicating stronger binding affinity with PET.

Table 1. Docking scores (binding energy, kcal/mol) of PET with dsPETase and mutants.

The plasmid map was constructed by snapgene software(Fig 8), and inserted into the expression vector pET28a by homologous recombination method, the recombinant plasmids were further transformed into E.Coli DH5α.

Fig 8. The plasmid map of pET28a-IsPETase

Build:

Fig. 9.The electrophoresis, plate Assay, and sequencing of dsPETase

The construction of the dsPETase plasmid employed an identical strategy to that of the IsPETase plasmid, with both based on the method of homologous recombination. As shown in Fig 9A, the dsPETase gene migrated between 500 and 1000 bp. This is consistent with its actual length of 816 bp, which was determined by sequencing. The linearized pET28a line is around 5000bp and its original length is 5329bp. We was transformed the pET28a-dsPETase plasmid into E.coli in the Fig 9C. In Fig 9D, sequencing of the plasmid confirmed that the inserted DNA fragment matched the target gene sequence, demonstrating successful ligation into the vector.

Fig 10. Agarose gel electrophoresis analysis of the mutant

We performed site-directed mutagenesis using PCR. As shown in Fig 10, the DNA fragments migrated between 5000 bp and 7500 bp, with all mutants (Arg47-Lys, Arg47-His, Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr) exhibiting a consistent length of 6190 bp. The plasmids were subsequently sent for commercial sequencing(Fig 11), and the results confirmed that the introduced mutations matched the expected designs.

Fig 11. The sequencing of the mutant

Test：

1. Protein Structure

1.1 Mutant Arg47-Lys and Arg47-His



Fig 12. Molecular interactions between the protein and the ligand within the binding pocket. Note: The protein structures of the Arg47-Lys and Arg47-His mutants are depicted in blue. The PET molecule is shown in orange.

The binding energies between the PET molecule and the mutant proteins Arg47-Lys and Arg47-His were −5.076 kcal/mol and −5.012 kcal/mol, respectively, indicating strong molecular interactions. As shown in Fig 12, the PET molecule interacts with the Arg47-Lys mutant primarily through hydrogen bonds and hydrophobic interactions. Hydrogen bonds were formed with LEU-70 and ALA-71, while hydrophobic interactions occurred with VAL-43, LEU-70, ALA-71, and ARG-68. In the Arg47-His mutant, the PET molecule was involved in hydrophobic interactions with the specific residues PHE-44, TYR-118, and THR-117 (Fig 12).

1.2 Mutant (Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr)



Fig 13. Molecular interactions between the protein and ligand within the binding pocket. The mutant protein structure is shown in blue, and the PET molecule is depicted in orange.

The binding energy between the PET molecule and the Gly8-Asn mutant protein was calculated to be −5.002 kcal/mol, indicating strong binding affinity. As shown in Fig 13, the PET molecule interacts with the Gly8-Asn mutant primarily through hydrogen bonding and hydrophobic interactions. Specifically, a hydrogen bond was formed with TYR-106, while hydrophobic interactions were observed with PHE-44, THR-117, and TYR-118.

The binding energy for the PET molecule with the Gly8-Cys mutant was −4.945 kcal/mol, demonstrating favorable binding interactions. Fig 14 shows that the PET molecule engages in hydrophobic interactions with LEU-103 and LYS-102, as well as electrostatic interactions with LYS-102 and ARG-99 in the Gly8-Cys mutant.



Fig 14. Molecular interactions between the protein and ligand within the binding pocket. The mutant protein structure is shown in blue, and the PET molecule is depicted in orange.

The binding energies between the PET molecule and the Gly8-Gln and Gly8-Ser mutant proteins were calculated to be −5.103 kcal/mol and −4.932 kcal/mol, respectively, indicating strong binding affinity. As shown in Fig 15, the PET molecule interacts with the Gly8-Gln mutant primarily through hydrogen bonding and hydrophobic interactions. Specifically, a hydrogen bond was formed with TYR-106, while hydrophobic interactions were observed with PHE-44, THR-117, and TYR-118. Fig 14 illustrates that the PET molecule engages in hydrophobic interactions with LEU-103 and LYS-102, as well as electrostatic interactions with LYS-102 and ARG-99 in the Gly8-Ser mutant.



Fig 15. Molecular interactions between the protein and ligand within the binding pocket. The mutant protein structure is shown in blue, and the PET molecule is depicted in orange.

The binding energies between the PET molecule and the Gly8-Thr and Gly8-Tyr mutant proteins were calculated to be −5.151 kcal/mol and −5.204 kcal/mol, respectively, indicating strong binding interactions. In Fig 15, the PET molecule engages in hydrophobic interactions with PHE-44, TYR-118, and THR-117 in the Gly8-Thr mutant. Fig 16 illustrates that the PET molecule interacts with the Gly8-Tyr mutant primarily through hydrogen bonding and hydrophobic interactions. Specifically, a hydrogen bond was formed with TYR-106, while hydrophobic interactions were observed with PHE-44 and TYR-118.



Fig 16.The growth curve of dsPETase and mutant

As shown in Figure 17, no significant difference was observed between the growth curves of the control group and the dsPETase and mutant strain, indicating robust bacterial growth.

2. Protein Expression



Fig 17. The SDS-PAGE of dsPETase and mutant. M:marker 1,2:rough protein; 3-5:Wash buffer 6-9:purified protein. DsPETase:31.5kDa

As shown in Figure 17A, lanes 1–2 exhibited diffuse bands in the crude protein samples, suggesting the presence of a substantial amount of non-specific proteins. Lanes 3–5, which served as blank controls, showed no detectable target band corresponding to dsPETase. In lanes 6–9, clear protein bands were observed within the molecular weight range of 25–35 kDa. These samples were purified by nickel-affinity chromatography, wherein Ni-NTA resin selectively captured His-tagged proteins, wash buffer removed non-specifically bound impurities, and the target protein was subsequently eluted under appropriate conditions. Figure 17B displays the protein expression profiles of various mutants. Band intensity and thickness reflect relative expression levels, with the Gly8-Ser mutant showing the darkest and thickest band, indicating its highest expression among all variants.

3. Determine the content of the product MHET

The chromatographic conditions were optimized using a Thermo Scientific C18 column (250 × 4.6 mm, 5 μm) with a mobile phase consisting of phosphate buffer (containing 5 mmol/L potassium dihydrogen phosphate and 0.04% phosphoric acid) and methanol in a ratio of 70:30 (v/v). According to the experimental protocol, standard solutions were serially diluted to appropriate concentrations to establish a calibration curve. The peak area was plotted on the Y-axis against the concentration of the reference standard on the X-axis, and regression analysis was performed to obtain the equation for each standard. A good linear relationship was observed within the concentration range of 1–50 mg/L (Table 2).

Table 2. The standard curve of MHET

Concentrations (mg/L)	Peak area
1	58.41312
2	130.59349
5	357.34796
10	697.24371
20	1393.50171
50	3568.67041

We incubated 500 mg of PET plastic substrates with IsPETase, dsPETase, and their variants at a final concentration of 1 mg/mL, then collected samples after 48 and 72 hours of degradation at room temperature. The peak area of MHET was measured by HPLC, and its concentration in the experimental samples was calculated based on the standard curve.

As shown in Fig 18A, MHET was detected in the experimental groups of IsPETase, dsPETase, and their mutants at 48-hour, whereas MHET was undetectable in the CK control group. Comparative analysis revealed no significant difference in MHET levels between IsPETase and dsPETase at the 48-hour. In contrast, all tested mutants—namely Arg47-Lys, Arg47-His, Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr—exhibited significantly higher MHET accumulation compared to dsPETase. Notably, the Gly8-Ser mutant generated markedly elevated MHET levels relative to the other mutants. Furthermore, there was no statistically significant difference in MHET production between the Arg47-Lys and Arg47-His variants.

In Fig 18B, MHET was detected in the experimental groups of IsPETase, dsPETase, and their mutants after 72 hours, while no MHET was observed in the CK control group. The MHET level produced by dsPETase was significantly higher than that of IsPETase at 72h. The MHET concentration of Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr mutants exhibited markedly higher than dsPETase. In contrast, no significant difference in MHET levels was observed among Arg47-Lys, Arg47-His, and dsPETase. These results suggest that Gly8 may represent a critical active site in dsPETase. Notably, the substitution of Gly8 with Ser significantly enhanced the degradation efficiency of PET.



Fig 18. MHET content of IsPETase, dsPETase, and mutantsArg47-Lys, Arg47-His, Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr) at 48 and 72 hours.

Learn:

In cycle 2, the plasmids for dsPETase and eight mutants (Arg47-Lys, Arg47-His, Gly8-Gln, Gly8-Asn, Gly8-Cys, Gly8-Ser, Gly8-Thr, and Gly8-Tyr) were successfully constructed and transformed into E. coli. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) analysis confirmed that the expressed proteins corresponded to the expected molecular sizes of dsPETase and all mutants, indicating their successful expression in E. coli. The experimental data from this cycle not only verified the ability of dsPETase and the mutants to degrade PET plastic but also enabled us to identify that the Gly8-Ser mutant exhibited higher enzymatic activity compared to the other mutants. These findings provide critical support for the subsequent dual-enzyme synergy strategy in cycle 3, making Gly8-Ser an indispensable component for the next phase.

Design:

Mono-2-hydroxyethyl terephthalate (MHET) is an intermediate with a competitive inhibitory effect in the degradation process of PET, which can be hydrolyzed from terephthalic acid (TPA) and ethylene glycol (EG) catalyzed by MHET degradation enzymes.

The genes encoding MHETase fused with 6×His-tag at its C-terminus were optimized and chemically synthesized. The plasmid map was constructed by snapgene software(Fig 19), and inserted into the expression vector pET28a by homologous recombination method, the recombinant plasmids pET28a- MHETase. were further transformed into E.Coli DH5α.

Fig 19. The plasmid map of pET28a- MHETase.

Build:

Fig 20.The electrophoresis, plate Assay, and sequencing of MHETase.

Note:the length of MHETase is 1818bp. the length of pET28a is 5324b

The agarose gel electrophoresis result in Fig 20 A and B showed a band of the MHETase gene at approximately 1500 bp and a linearized plasmid band near the 3000 bp marker. After ligation, the recombinant plasmids were transformed into E. coli DH5α competent cells for amplification in the Fig 20C. The subsequent sequencing analysis confirmed the successful insertion of the target gene MHETase into the vector(Fig 20D).

Test：

1. Protein expression



Fig 21.The growth curve and SDS-PAGE of IsPETase. Note:The size of MHETase is 64kDa.

To express the target protein, the plasmid was transformed into E. coli BL21(DE3) cells and induced with 1 mM IPTG, followed by protein extraction and purification. In Fig 21A, no significant difference was observed between the growth curves of the control group and the dsPETase and mutant strain, indicating robust bacterial growth. The SDS-PAGE analysis showed that the purified MHETase protein was approximately 63 kDa, which is consistent with its actual molecular size(Fig 21 B).

2. Scanning electron microscopy (SEM)



Fig. 22. Results of scanning electron microscopy (SEM) of PET. A: control group; B:IsPETase+MHETase; C:DsPETase+MHETase; D:Gly8-Ser+MHETase

Fig 22A shows the PET film from the control group without enzyme addition, exhibiting a smooth surface without cracks or pores. Fig 22B presents the experimental group treated with IsPETase and MHETase, where faint fissures observed on the film surface indicate initial enzymatic activity, though the degradation effect remains suboptimal. In Fig22 C, the sample co-treated with dsPETase and MHETase demonstrates more distinct fissure structures, suggesting enhanced degradation efficacy compared to the IsPETase group. Figure 22 D displays the Gly8-Ser variant + MHETase group. Numerous pronounced fissures can be observed on the film surface, demonstrating the superior degradation performance of the Gly8-Ser variant.

3. HPLC determination of concentration TPA and EG

The experimental instrument parameters were adjusted as follows: the column was a Thermo Fisher C18 (250 mm × 4.6 mm, 5 μm), with a column temperature of 30°C, flow rate of 1 mL/min, and a mobile phase consisting of phosphate buffer solution (containing 5 mmol/L potassium dihydrogen phosphate and 0.04% phosphoric acid) and methanol at a ratio of 70:30.

According to the experimental protocol, the standard was diluted to appropriate concentrations to establish a calibration curve. Using the peak area as the ordinate (Y) and the mass concentration of the reference standard as the abscissa (X), regression analysis was performed to derive the equation for each standard. A good linear relationship was demonstrated within the concentration range of 1–50 mg/L. The tested concentrations and corresponding calibration curve are presented in Table 3.

Table 3. The standard curve of TPA

Concentrations (mg/L)	Peak area
1	85.65078
2	204.93451
5	584.46924
20	2220.17896
50	6006.84082

We incubated 1 g PET plastic with 1 mg/mL IsPETase, dsPETase, and the Gly8-Ser variant, along with MHETase at a final concentration of 1 mg/mL. Under the catalytic action of PETase and MHETase, PET can be degraded to TPA, which was quantified using HPLC. As shown in Fig 23, the control group (pET28a vector) showed no peak at 10–12 minutes, whereas distinct peaks were observed for the IsPETase + MHETase, dsPETase + MHETase, and Gly8-Ser variant + MHETase groups.



Fig. 23. Chromatographic peak of TPA

As illustrated in Figure 24, no TPA was detected in the control (CK) group. The TPA concentrations produced in dsPETase + MHETase and Gly8-Ser + MHETase were significantly higher than IsPETase + MHETase. Furthermore, the TPA concentrations from the Gly8-Ser+ MHETase combination was markedly higher than that of dsPETase + MHETase. These results indicate that the degradation efficiency of dsPETase + MHETase exceeds that of IsPETase + MHETase, and the Gly8-Ser variant in combination with MHETase demonstrated superior performance compared to all other experimental groups. This evidence confirms that site-directed mutagenesis enhanced the enzymatic activity of PETase, thereby significantly improving the overall degradation efficiency.



Fig. 24. The concentration of TPA of CK, IsPETase + MHETase, dsPETase + MHETase, and Gly8-Ser+ MHETase

EG (Ethylene Glycol) was qualitatively and quantitatively analyzed in complex mixtures using Gas Chromatography-Mass Spectrometry (GC-MS). The instrument parameters were set as follows:

Gas Chromatograph: Agilent GC 8860 equipped with CTC PAL3 autosampler

Mass Spectrometer: Agilent 5977B MSD

Column: Agilent DB-WAX (30 m × 0.25 μm)

Carrier Gas: High-purity helium (≥99.99999%)

Chromatographic conditions:

Detector temperature: 240°C

Injector temperature: 220°C

Flow rate: 1.0 mL/min

Temperature gradient: Initial temperature 80°C (hold 5 min), then ramp at 5°C/min to 240°C (hold 1 min).



Fig. 25. The chromatographic peak, ratio, mass scharge ratio of EG.

As shown in Fig 25, the detection results of ethylene glycol (EG) in the samples were obtained using high-performance liquid chromatography (HPLC) and mass spectrometry (MS). The control group exhibited no detectable peaks, indicating the absence of EG in the sample. Distinct chromatographic peaks were observed in the experimental groups containing IsPETase + MHETase, dsPETase + MHETase, and the Gly8-Ser + MHETase.The concentration of EG in the samples was calculated based on the chromatographic peak areas using the standard curve equation y = 6.306x + 363.8. As shown in Figure 26 A, the calibration curve exhibited an excellent fit with a coefficient of determination (R²) greater than 0.99.

In Fig 26B, the EG concentration obtained with dsPETase + MHETase was significantly higher than that with IsPETase + MHETase. Notably, the Gly8-Ser variant + MHETase combination yielded a substantially greater EG concentration compared to both the dsPETase and IsPETase. These results indicate that the Gly8-to-Ser mutation at position 8 in dsPETase enhances enzymatic activity, significantly improving the conversion efficiency of EG.



Fig 26. The concentration of EG of CK, IsPETase + MHETase, dsPETase + MHETase, and Gly8-Ser+MHETase

Learn:

Through this study, we demonstrate the feasibility of employing a dual-enzyme system—comprising dsPETase and MHETase—for synergistic degradation of polyethylene terephthalate (PET), enabling a closed-loop recycling process. Our work primarily focused on enhancing the depolymerization efficiency via structure-guided engineering of dsPETase. Using molecular docking, we predicted key residues within the enzyme’s active site and conducted site-directed mutagenesis at positions Arg47 and Gly8, substituting them with amino acids of similar properties to minimize functional disruption.

Experimental results revealed that the Gly8-Ser mutation significantly improved PET degradation efficiency, whereas the Arg47 variant showed no notable effect. Further optimization of catalytic activity may be achieved through additional rational mutagenesis at predicted active sites. Moreover, the combined action of the Gly8-Ser and MHETase synergistically catalyzed the complete decomposition of PET into its monomers, terephthalic acid (TPA) and ethylene glycol (EG), which can be reused as raw materials for plastic synthesis.

In summary, this study proposes a novel enzymatic strategy for sustainable plastic degradation and recycling. These findings provide a theoretical foundation for the development of biocatalytic recycling processes and highlight potential applications in the bioremediation of plastic pollution.

Guo, Z., Li, Y., Wang, M., & Ma, D. (2025). Catalytic upcycling of PET: From waste to chemicals and degradable polymers. Accounts of Chemical Research.

KANELLI M, VASILAKOS S, NIKOLAIVITS E, LADAS S, CHRISTAKOPOULOS P, TOPAKAS E. Surface modification of poly(ethylene terephthalate) (PET) fibers by a cutinase from Fusarium oxysporum[J]. Process Biochemistry, 2015, 50(11): 1885-1892. DOI:10.1016/j.procbio.2015.08.013

V T ,M C T ,A G , et al.An engineered PET depolymerase to break down and recycle plastic bottles.[J].Nature,2020,580(7802):216-219.

Tournier, V., et al. (2023). Enzymatic recycling of PET plastic: From discovery to industrial implementation. Nature Reviews Chemistry, 7(12),

YOSHIDA S, HIRAGA K, TAKEHANA T, TANIGUCHI I, YAMAJI H, MAEDA Y, TOYOHARA K, MIYAMOTO K, KIMURA Y, ODA K. A bacterium that degrades and assimilates poly(ethylene terephthalate). Science, 2016, 351(6278): 1196-1199. DOI:10.1126/science.aad6359