ACS Publications. Most Trusted. Most Cited. Most Read
Particle Size Reduction of Poly(ethylene terephthalate) Increases the Rate of Enzymatic Depolymerization But Does Not Increase the Overall Conversion Extent
My Activity

Figure 1Loading Img
  • Open Access
Research Article

Particle Size Reduction of Poly(ethylene terephthalate) Increases the Rate of Enzymatic Depolymerization But Does Not Increase the Overall Conversion Extent
Click to copy article linkArticle link copied!

  • Richard K. Brizendine
    Richard K. Brizendine
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    BOTTLE Consortium, Golden, Colorado 80401, United States
  • Erika Erickson
    Erika Erickson
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    BOTTLE Consortium, Golden, Colorado 80401, United States
  • Stefan J. Haugen
    Stefan J. Haugen
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
  • Kelsey J. Ramirez
    Kelsey J. Ramirez
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    BOTTLE Consortium, Golden, Colorado 80401, United States
  • Joel Miscall
    Joel Miscall
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    BOTTLE Consortium, Golden, Colorado 80401, United States
    More by Joel Miscall
  • Davinia Salvachúa
    Davinia Salvachúa
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
  • Andrew R. Pickford
    Andrew R. Pickford
    BOTTLE Consortium, Golden, Colorado 80401, United States
    Centre for Enzyme Innovation, School of Biological Sciences, Institute of Biological and Biomedical Sciences, University of Portsmouth, Portsmouth PO1 2DY, U.K.
  • Margaret J. Sobkowicz
    Margaret J. Sobkowicz
    Department of Plastics Engineering, University of Massachusetts Lowell, Lowell, Massachusetts 01854, United States
  • John E. McGeehan*
    John E. McGeehan
    BOTTLE Consortium, Golden, Colorado 80401, United States
    Centre for Enzyme Innovation, School of Biological Sciences, Institute of Biological and Biomedical Sciences, University of Portsmouth, Portsmouth PO1 2DY, U.K.
    *Email: [email protected]
  • Gregg T. Beckham*
    Gregg T. Beckham
    Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    BOTTLE Consortium, Golden, Colorado 80401, United States
    *Email: [email protected]
Open PDFSupporting Information (1)

ACS Sustainable Chemistry & Engineering

Cite this: ACS Sustainable Chem. Eng. 2022, 10, 28, 9131–9140
Click to copy citationCitation copied!
https://doi.org/10.1021/acssuschemeng.2c01961
Published July 7, 2022

Copyright © 2022 The Authors. Published by American Chemical Society. This publication is licensed under

CC-BY 4.0 .

Abstract

Click to copy section linkSection link copied!

Enzymatic depolymerization of poly(ethylene terephthalate) (PET) has emerged as a potential method for PET recycling, but extensive thermomechanical preprocessing to reduce both the crystallinity and particle size of PET is often conducted, which is costly and energy-intensive. In the current work, we use high-crystallinity PET (HC-PET) and low-crystallinity cryomilled PET (CM-PET) with three distinct particle size distributions to investigate the effect of PET particle size and crystallinity on the performance of a variant of the leaf compost-cutinase enzyme (LCC-ICCG). We show that LCC-ICCG hydrolyzes PET, resulting in the accumulation of terephthalic acid and, interestingly, also releases significant amount of mono(2-hydroxyethyl)terephthalate. Particle size reduction of PET increased the maximum rate of reaction for HC-PET, while the maximum hydrolysis rate for CM-PET was not significantly different across particle sizes. For both substrates, however, we show that particle size reduction has little effect on the overall conversion extent. Specifically, the CM-PET film was converted to 99 ± 0.2% mass loss within 48 h, while the HC-PET powder reached only 23.5 ± 0.0% conversion in 144 h. Overall, these results suggest that amorphization of PET is a necessary pretreatment step for enzymatic PET recycling using the LCC-ICCG enzyme but that particle size reduction may not be required.

This publication is licensed under

CC-BY 4.0 .
  • cc licence
  • by licence
Copyright © 2022 The Authors. Published by American Chemical Society

Synopsis

PET particle size reduction increases the rate but not the extent of conversion by a PET-degrading enzyme.

Introduction

Click to copy section linkSection link copied!

Plastics are necessary in today’s world, but most plastics are not recycled and instead are discarded in landfills or accumulate in the environment. (1−3) Poly(ethylene terephthalate) (PET), with ∼65 million metric tons produced annually, (4−6) is the second most abundantly produced plastic, behind only polyethylene. Currently, the most common type of PET recycling is thermomechanical, which often results in recycled polymers with less desirable mechanical properties than those produced from virgin terephthalic acid (TPA) and ethylene glycol, (7) which are the monomers used for PET synthesis. Therefore, chemical recycling technologies that can depolymerize PET to its constituent monomers are of keen interest to regenerate virgin-quality PET. (8)
Enzymatic recycling is an emerging chemical recycling strategy for PET due to the relatively low temperature and pressure required, as well as high selectivity in the enzymatic reactions, possibly enabling precise PET depolymerization within complex mixed waste streams that have been previously excluded from recycling processes. (9−11) Many PET-degrading enzymes have been previously reported and characterized, (12−25) but most show limited activity on PET and are therefore likely not directly suitable for industrial use. There have recently been many efforts to enhance the thermostability and catalytic activity of several PET hydrolases to this end. (26−46)
To date, some of the highest depolymerization extents of PET were reported by Tournier et al. (39) The authors reported an engineered variant of leaf compost-cutinase (LCC) (17,19) capable of degrading PET to >90% within 10 h. While the authors concluded that this performance is sufficient for industrial-scale recycling, the PET used in their study was extensively preprocessed to reduce long-range order and crystallinity (amorphization) and reduce particle size (micronization) prior to enzymatic degradation. These are fundamental process parameters, given that techno-economic analysis (TEA) suggests that mechanical preprocessing of PET is a main contributor to cost, energy consumption, and greenhouse gas emissions of the process. (47) Specifically, the analysis suggests that by eliminating mechanical pretreatment, the process electricity usage decreases by 67%, the overall process energy reduces by almost 50%, and the minimum selling price of the recovered TPA is reduced by $0.24/kg. (47) However, to our knowledge, little work has been reported on the effect of amorphization and micronization of PET with these recently developed enzymes. It is well accepted that many reported PET-hydrolyzing enzymes work better on amorphous PET, (15,38,48) with Ideonella sakaiensis PETase (IsPETase) and several recently reported variants thereof being possible exceptions. (22,34,40,49−51) Moreover, studies from Castro et al. and Gamerith et al. have shown that reducing the PET particle size increased the conversion rate and the overall extent of conversion; (52−54) however, the enzymes used have not been reported to achieve the same conversion extents as the current engineered LCC variant. (39)
In this study, we investigated the effect of particle size reduction and crystallinity on the performance of PET depolymerization by the LCC-ICCG variant. (39) We prepared PET particles with low and high crystallinities across three particle size ranges each, and the PET hydrolysis performance of LCC-ICCG was tested on these substrates at varying solid and enzyme loadings. We used the inverse Michaelis–Menten (invMM) analysis proposed by Westh et al. to compare the enzyme kinetics across the PET substrates. (55−57) Additionally, we performed reactions in bioreactors to investigate the enzyme performance in a pH-controlled environment. Our results suggest that decreasing the particle size of PET increases the initial rate of the reaction but has little effect on the overall conversion extent. Instead, the results show that crystallinity of the PET substrate is the key driver of enzymatic PET degradation with this enzyme system.

Materials and Methods

Click to copy section linkSection link copied!

Reagents, Stocks, and Buffers

All chemicals were obtained from Sigma-Aldrich at the highest available grade of purity and used as supplied, unless otherwise noted. Buffers were made using ultrapure water. Equilibration buffer: 20 mM Tris, pH 8, 300 mM NaCl, and 10 mM imidazole. Elution buffer: 20 mM Tris, pH 8, 300 mM NaCl, and 500 mM imidazole. Assay buffer: 100 mM NaPi, pH 8.

Preparation and Characterization of PET Particles

Amorphous PET particles (which we denote CM-PET) were prepared by cryomilling a 0.25 mm thick amorphous PET film (A-PET, Goodfellow, ES301445). The film was cut into squares with 0.5 × 0.5 cm dimensions and cryomilled in 1–2 g aliquots in stainless-steel vials using a Freezer Mill 6770 (Spex SamplePrep) and ground for a total of 40 min using the following grinding cycle: 4 min grinding, 2 min cooling, with 10 cycles in total. High-crystallinity PET particles (denoted HC-PET) were sourced from Goodfellow (ES306031).
To separate the particles by size, the PET particles were wet sieved using two sieve sizes: 250 and 125 μm sieves (Prüfsieb). Briefly, 5–10 g of PET powder was placed in the 250 μm sieve and ddH2O was slowly run over the top. The filtrate was collected until no PET was observed in the water (usually 8–10 L). This process was repeated with the 125 μm sieve. The PET that went through the 125 μm sieve was collected by filtering the solution using Whatman grade 5 filter paper (Cytiva) and a Büchner funnel. The PET remaining in the 250 μm sieve was termed the “250 μm fraction”, the PET remaining in the 125 μm sieve was similarly termed the “125 μm fraction”, and the PET that went through the 125 μm sieve was denoted as the “sub125 μm fraction”. All PET particles were dried at 40 °C for 2 days under vacuum.
The particle size distributions of each sieve fraction were determined using dark-field stereomicroscopy. PET particles were suspended in 1% (w/v) sodium dodecyl sulfate and spread over a glass slide (Fisher Scientific) at a concentration that reduced contact between individual particles (∼500 μg mL–1). The 250 and 125 μm sieve fractions were imaged at 2× magnification, and the sub125 μm sieve fraction was imaged at 4× magnification on a Nikon SMZ1500 stereomicroscope equipped with a Nikon Ds-Fi1 camera. Particle sizes were obtained by analysis with a custom ImageJ macro (NIH). Briefly, individual particles were segmented by making the image binary and using Analyze Particles function. Particles were fit to an ellipse, and the surface area and volume were estimated by assuming the height of the particle was the average of the short and long axes of the ellipse. The mass of the particles was estimated using the density of PET, 1.38 g cm–3. The specific surface area was then calculated as the estimated sum of surface area over the sum of the mass of all analyzed particles.
Detailed imaging of individual PET particles was performed on a Nikon Eclipse E800 using either 10× 0.45 NA or 20× 0.75 NA PlanApo Nikon objectives, dark-field illumination, and a SPOT RTKE CCD camera (Diagnostic Instruments). PET particles were sprinkled onto a glass slide and distributed to isolate the particles. Image slices were collected at varying depths of field by manually changing the focus from the slide surface to the top of the particle and capturing images every ∼2–5 μm. The images were stacked using the Focus Merge function in Affinity Photo (Serif).
The crystallinity of all PET substrates was determined by differential scanning calorimetry (DSC) using a Q2000 DSC (TA Instruments) on 2–8 mg of PET samples placed in hermetically sealed aluminum pans. The samples were analyzed from 0 to 300 °C at a rate of 10 °C min–1. The glass-transition temperature (Tg), heat of melting (ΔHm), and heat of cold crystallization (ΔHcc) were determined with Universal Analysis (TA Instruments). Percent crystallinity was calculated using the following equation, where ΔHm° is the reference heat of melting for PET = 140.1 J g–1
(1)
The weight-average molar mass (Mw) and number-average molar mass (Mn) were determined by gel permeation chromatography (GPC) using an Agilent 1260 Infinity II LC system, which consisted of a 1260 Iso pump module, 1260 vial sampler module, and a 1260 Multicolumn Thermosat module. Three PL HFIPgel 250 × 4.6 mm columns (Agilent, PL1514-5900HFIP) attached in series were used for analysis, with a matching guard column attached. Samples were prepared in 0.22 μm filtered hexafluoroisopropanol (HFIP) at a concentration of 2 mg mL–1 and filtered through a 0.22 μm filter. The operating conditions included HFIP as the carrier solvent, a flow rate of 0.4 mL min–1, column temperature set to 40 °C, and a sample injection of 100 μL. Detectors consisted of a miniDAWN TREOS multiangle light-scattering detector (Wyatt Technology) used in combination with an Optilab T-rEX refractive index detector (Wyatt Technology). Wyatt Technologies Astra 7.2 software was used to analyze the data.

Expression and Purification of the LCC-ICCG Enzyme

The LCC-ICCG enzyme was expressed and purified similar to the method reported by Tournier et al. (39) The LCC-ICCG DNA was synthesized (Twist Bioscience) and cloned into pET-21b(+) (EMD Biosciences). The plasmid was then transformed into OverExpress Escherichia coli C41 (DE3) (Lucigen) cells according to the manufacturer’s protocol, plated on lysogeny broth (LB)-agar plates containing 100 μg mL–1 ampicillin (Amp), and incubated at 37 °C overnight. Single colonies from transformation were inoculated into a starter culture of LB liquid media containing 100 μg mL–1 Amp and grown at 37 °C with shaking at 250 rpm overnight. The starter culture was inoculated at a 100-fold dilution in 2× YT media (10 g of yeast extract, 16 g of tryptone, and 10 g of NaCl per 1 L media) containing 100 μg mL–1 Amp and grown at 37 °C to OD600 = 0.6–0.8. Protein expression was induced by the addition of isopropyl β-d-1-thiogalactopyranoside (IPTG) to a final concentration of 1 mM. Cells were maintained at 20 °C and 225 rpm for 18–24 h following IPTG induction, harvested by centrifugation, and stored at −80 °C until purification. Harvested cells were resuspended in lysis buffer containing 1 mg mL–1 lysozyme and 50 μg mL–1 DNAase and incubated on ice for 10 min. Cells were homogenized on ice using a sonicator (QSonica Q700) set to 50% amplitude in 10 s bursts, followed by 15–20 s cooling breaks. Total sonication time was 2 min. The lysate was clarified by centrifugation at 40,000g for 40 min at 4 °C. Clarified lysate was then applied to a 5 or 25 mL HisTrap HP (Cytiva) column using an ÄKTA Pure chromatography system (Cytiva) and eluted with elution buffer over a 2 CV gradient. Fractions containing the protein were dialyzed overnight against 20 mM Tris, pH 8, 300 mM NaCl at room temperature. The sample was then clarified by filtration through a 0.45 μm polyvinylidene fluoride syringe filter (Thermo Scientific). The protein was then further purified by size-exclusion chromatography with HiLoad 16/60 Superdex 75 pg column (Cytiva). The enzyme could also be stored for several days at 4 °C, which caused most remaining impurities to precipitate, which were then removed by centrifugation or filtration. The enzyme was stored for up to 6 months at 4 °C with no detectable loss in activity.

Enzymatic Hydrolysis Experiments

Small-scale 1.5 mL volume reactions were performed in triplicate in 2 mL polypropylene Eppendorf tubes containing assay buffer and varying masses of PET substrate (0.5–10 mg mL–1). Reaction tubes were equilibrated to 65 °C in a heat block (Thermo Scientific) for 10 min, and the reaction was started by the addition of varying concentrations of LCC-ICCG (30 nM to 5 μM final). Controls were performed with no added enzyme. Timepoints of 125 μL were removed with rapid pipetting to suspend the PET powder and immediately quenched into an equal volume of methanol. Samples were filtered through a 96-well, 0.25 μm filter plate (EMD Millipore, MSGVN22) using a vacuum manifold (Pall, 5017) into a 300 μL collection plate (Agilent, 5043-9312) and sealed with a silicone mat (Agilent, 5043-9317).
Large-scale reactions were performed at 1 L scale in duplicate in 3 L glass bioreactors (Applikon Biotechnology), which included two Rushton impellers in the stirrer shaft below the 1 L line. The gas-sparging arm and baffles were removed. Three different substrates were used: A-PET (Goodfellow, ES301445) cut into ∼1 × 1 cm squares, HC-PET that had not been separated by sieving (Goodfellow, ES306031), and residual PET fines from a bottle-to-bottle recycling plant kindly provided by Western Container Corporation (WC-PET). Briefly, 100 g of PET substrate (73 mL volume based on density of PET) was added to 893 mL of assay buffer and equilibrated to 65 °C with stirring at 250 rpm. The reaction was initiated by the addition of 35 mL of 8.6 mg mL–1 LCC-ICCG for a final enzyme loading of 3 mg g–1 PET (10.5 μM LCC-ICCG). No-enzyme controls were performed in 250 mL of assay buffer with 25 g of PET substrate in 250 mL glass jars (Corning), maintained at 65 °C in a MaxQ 6000 shaking incubator (Thermo Scientific) set at 250 rpm. Reactions proceeded for 144 h and were maintained at pH 8 with 4 N NaOH addition using a peristaltic pump controlled by an in-control module (Applikon Biotechnology). Control reactions did not require pH control since abiotic PET hydrolysis was very low or undetectable. Sample volumes of 1 mL were removed at designated time points, quenched, stored, and filtered as described above for the small-scale reactions. At the end of the reaction, the remaining A-PET was collected by filtration through Whatman grade 2 filter paper (Cytiva) and a Büchner funnel, while the remaining HC-PET and WC-PET were collected by filtration through 0.45 μm jar-top filter units (Thermo Scientific, 169-0045). The filters were preweighed, and after the PET was collected, the filters with PET were dried for 3 days at 50 °C under vacuum before the final mass of residual PET was calculated.

Ultrahigh-Performance Liquid Chromatography Analysis of Reaction Timepoints

Standards of bis(2-hydroxyethyl)terephthalate (BHET) and TPA were acquired from Sigma-Aldrich. Mono(2-hydroxyethyl)terephthalate (MHET) was prepared as described previously. (49) Quantitation of BHET, MHET, and TPA was performed using ultra-high-performance liquid chromatography on an Infinity II 1290 system (Agilent Technologies), as described previously. (58)

Determination of Initial Rates and Kinetic Analysis

Initial rates of the reactions were determined by linear fits to the sum of the monomers released for each timepoint up to the first 5 h of the reaction. The invMM equation was used as described previously. (57) All linear and invMM fits were performed with Origin Pro 2021 (OriginLab Corporation). MHET autohydrolysis rates were fit to a single-step rate equation using KinTek Explorer (KinTek Corporation).

Results and Discussion

Click to copy section linkSection link copied!

Characterization of PET Substrates

Figure 1 shows particle size distributions of sieved PET substrates. The CM-PET particle (Figure 1A–C) size distributions are overall very similar to the HC-PET size distributions (Figure 1D,E), resulting in similar specific surface areas across the three sieve fractions and substrates (Table 1). Despite similar particle size distributions, the two sets of particle preparations exhibit different morphologies when examined with dark-field microscopy. The CM-PET particles display a variety of morphologies, including particles that appear shredded or torn (Figure S1A,C) and others that appear to be uniform, solid particles (Figure S1B). In contrast, the HC-PET particles are generally more consistently solid particles (Figure S1D–F). Interestingly, many of the HC-PET particles also exhibit complex morphological traits, including long offshoots from the main body of the particle (Figure S1D,E).
There are also substantial differences in the crystallinity of these two PET substrates. While the CM-PET particles originated from the LC-PET film (∼4% crystallinity, Table 1), the cryomilling process had a measurable effect on the crystallinity of the resulting particles since the crystallinity of the PET after cryomilling was higher than that of the starting material (Table 1 and Figure S2). Since the PET was ground at cryogenic temperatures, this increase is likely not due to heat-induced crystallization but instead could be attributed to stress-induced crystallization. (59−62) Some variation in the crystallinity of the sieve fractions was observed, with all samples having crystallinity between 7 and 15% (Table 1). A possible explanation for the variability could be artifacts of the DSC measurement, which can be variable and can tend to overestimate the crystallinity of amorphous PET. (63) Cryomilling of the PET film did not significantly alter the molecular-weight distributions as all three sieve fractions were similar to the starting material (Table 1). The HC-PET particles exhibit much higher crystallinity than the CM-PET and were more consistent across the sieve fractions, with all of them in the 32.5–35.7% crystallinity range (Table 1 and Figure S3). There was also no difference in the molecular weight distributions across the sieve fractions of the HC-PET particles (Table 1).

Effect of Particle Size and Crystallinity on Enzymatic Degradation of PET

To determine the effect of particle size on enzymatic degradation of PET, reactions were performed with different PET substrates. Figure 2, Dataset S1 shows representative time courses from small-scale depolymerization experiments conducted with 10 mg mL–1 PET and 1 μM LCC-ICCG. For the CM-PET (Figure 2A–C), the primary monomers released were TPA and MHET, with the MHET eventually converting to TPA over the course of the reaction. The monomer concentration eventually reached ∼70 mM. The HC-PET generally followed the same trend; however, the concentration of the monomers was overall much lower, reaching only ∼20–30 mM after 72 h (Figure 2D–F). Interestingly, for both the CM-PET and HC-PET particles, the MHET concentration appeared to be affected by the size of the particles. MHET reached the highest concentrations in the reactions with the sub125 μm PET particles for both the CM-PET and HC-PET particles.
To examine the transient nature of MHET accumulation under these reaction conditions, we evaluated whether LCC-ICCG could catalyze MHET hydrolysis. Incubation with MHET and LCC-ICCG showed that the enzyme did not hydrolyze the substrate faster than a control reaction with no enzyme (Figure S4, Dataset S2). However, MHET spontaneously hydrolyzes under these conditions at a rate of 0.082 h–1 (Figure S4). This result indicates that LCC-ICCG can convert PET to both MHET and TPA and that the MHET slowly hydrolyzes to TPA explaining the transient accumulation of MHET in the reactions with PET.

Figure 1

Figure 1. Size distributions of CM-PET and HC-PET particles. Histograms indicate the volume percentage as a function of Feret’s minimum diameter of CM-PET (blue) and HC-PET (red) substrates. (A,D) 250, (B,E) 125, and (C,F) sub125 μm sieve fractions. Inset graphs show the expanded view of the data.

Table 1. Characterization of PET Substratesa
PET substratesourcesieve fraction (μm)specific SA (m2 g–1)Tg (°C)% crystallinityMn (kDa)Mw (kDa)
LC-PETlow-crystallinity PET film, Goodfellow ES301445N/AN/A75.7 ± 0.64.2 ± 219.7 ± 8.333.8 ± 6.8
CM-PETcryomilled LC-PET, from Goodfellow ES3014452500.0179.0 ± 0.811.0 ± 1.820.9 ± 7.534.9 ± 6.7
  1250.0274.2 ± 0.37.6 ± 119.3 ± 8.634.8 ± 6.5
  sub1250.1269.4 ± 0.714.3 ± 2.320.3 ± 4.934.5 ± 6.4
HC-PETsemicrystalline powder, Goodfellow ES3060312500.01276.1 ± 0.335.7 ± 3.819.9 ± 6.432.5 ± 7.1
  1250.02276.6 ± 0.633.0 ± 2.119.2 ± 5.733.0 ± 6.3
  sub1250.0676.8 ± 0.632.5 ± 1.720.1 ± 7.232.1 ± 6.9
WC-PETfines supplied by Western Container CorporationN/AN/A70.0 ± 0.537.3 ± 2.526.7 ± 8.443.5 ± 6.6
a

Specific surface area calculated from stereomicroscopy, DSC data, and GPC data for all PET substrates. ± indicates SD, n = 3.

To determine the effect of enzyme concentration on overall performance, we repeated the experiments shown in Figure 2 at varying enzyme concentrations (from 0.03 to 1.0 μM) and compared the concentration of monomers released after 72 h. Figure 3, Dataset S3 shows the results of these experiments. Generally, the highest extents of conversion were reached at the highest enzyme concentrations for both CM-PET and HC-PET, with very little overall difference observed between the different particle sizes. The CM-PET reached total monomer concentrations of ∼70–80 mM, while the HC-PET product yields were ∼3-fold lower at ∼20–30 mM.
To further examine how particle size and crystallinity affect the initial rate and overall performance of the LCC-ICCG enzyme, we followed the approach described by Bååth et al. (57) In this approach, two different experiments are conducted: one where the PET loadings are varied with a constant enzyme concentration [conventional Michaelis–Menten (MM) approach] and one where the enzyme concentration is varied while holding the PET loading constant (invMM approach). (15,55,64) Analysis of these two experiments can provide insights into numbers of available binding sites for the enzyme on the PET surface as well as conventional specificity constants. Unfortunately, at the low enzyme concentrations and PET loadings required for conventional MM kinetics, the reaction profiles were not linear. Instead, there was a significant lag (Figure S5, Dataset S4), which makes determining the initial rates challenging and likely inaccurate. Thus, we only report the results of invMM analysis here.
Figure 4, Dataset S5 shows the invMM analysis (55,56) for the enzymatic hydrolysis of CM-PET (Figure 4A) and HC-PET particles (Figure 4B). The parameters resulting from the fits of the invMM equation to the data are shown in Table 2. The CM-PET particles exhibit similar invKm values and nearly identical invVmax. The largest difference between the small and large particles was observed in the HC-PET, with the sub125 μm particles reaching a invVmax approximately double that of the 250 μm sieve fraction. Generally, the smaller particle sizes resulted in higher initial rates than the larger particles. However, for the CM-PET particles, as the enzyme concentration increases, the rate of the reaction for all particles converges, whereas for the HC-PET, the initial rates were still notably different even at the high enzyme concentrations.
We also investigated the effect of PET particle size and crystallinity in 1 L scale reactions with pH control, shown in Figure 5, Dataset S6. We tested three different PET substrates: (1) A-PET cut into ∼1 × 1 cm squares (Figure 5A,D), (2) HC-PET (Figure 5B,E), and (3) WC-PET (Figure 5C,F). The A-PET cut into ∼1 × 1 cm squares performed the best, reaching 99 ± 0.3% conversion by mass loss and 91.3 ± 1.8% measured by HPLC analysis after ∼48 h (Figure 5D). In contrast, the higher-crystallinity powders HC-PET and WC-PET reached much lower conversion extents, only reaching 23.5 ± 0.0 and 24.6 ± 0.8% degradation by mass loss, respectively, and 19.2 ± 0.9 and 24.0 ± 0.1% as measured by HPLC after 144 h (Figure 5E,F). Note the error for these numbers is the absolute difference between duplicate experiments.

Figure 2

Figure 2. Monomer release as a function of time for 10 mg mL–1 PET particles with 1 μM LCC-ICCG. Monomers present in the reaction were analyzed by HPLC. (A) 250 μm CM-PET, (B) 125 μm CM-PET, (C) sub125 μm CM-PET, (D) 250 μm HC-PET, (E) 125 μm HC-PET, and (F) sub125 μm HC-PET. Reactions were performed in triplicate and errors bars represent the standard deviation. The data shown in this figure are provided in Dataset S1.

Generally, the monomer release profiles from the 1 L scale reactions were similar to those from the small-scale reactions. TPA was the primary product released over time for all substrates with MHET released in a similarly transient manner as the small-scale reactions. The MHET reached similar concentrations (∼10–12 mM) for all substrates tested in the bioreactors. Similarly, the LC-PET substrate (LC-PET, Figure 5A) generated significantly higher overall concentrations of monomers upon enzyme treatment, ∼450 mM, when compared to the higher-crystallinity HC-PET (Figure 5B) and WC-PET (Figure 5C), which each released ∼100–120 mM total monomers.
The goal of this study was to investigate the effect of PET particle size and crystallinity on the enzymatic degradation of PET with LCC-ICCG. To that end, we generated PET particles with similar sizes and surface areas but with low and high crystallinity (Figure 1 and Table 1). In small-scale tests, we observed that the sub125 μm sieve fraction particles generally achieved the highest extents of conversion during the initial 2–5 h of the reaction, suggesting that reducing the particle size has the largest effect on initial reaction rates (Figure 2). Interestingly, this observation did not necessarily translate to increased overall conversion extents since all particle sizes of a given PET substrate generally reached the same extent of conversion by the end of the reaction (Figure 3). Instead, crystallinity of the substrate appears to be the main driver for overall performance since the CM-PET (7–15% crystallinity) reached ∼3-fold higher conversion extents than the HC-PET (33–35% crystallinity). These data are consistent with previous studies using various PET hydrolase enzymes and PET substrates, which also show that as the crystallinity of the PET substrate increases, the degradation performance decreases. (15,38,48) These results highlight the need for enzymes that work well on high-crystallinity PET to reduce the necessity of costly and energy-intensive pretreatment. (4,50)
While overall depolymerization extent appeared to be largely unaffected by reducing the particle size of the PET, the initial rate of the reaction was affected. We investigated this by varying the enzyme concentration with the PET particle solid loading held constant at 10 mg mL–1 and using an invMM analysis (Figure 4). The results of this experiment were notably different between the two different PET crystallinities. First, the HC-PET particles across all size fractions had a lower invVmax than the corresponding CM-PET particles (Table 2). Second, comparing across the various particle sizes for each substrate, we found that the HC-PET particles with the smallest size uniformly exhibited the highest initial rate. This was not true for the CM-PET where the rates eventually converged with higher enzyme concentrations. If we assume that invVmax is governed by kcat, (57) then these data suggest that similar amounts of enzyme can bind to the various particle sizes of the CM-PET, resulting in similar maximal rates. However, for the HC-PET, the invVmax is higher for the sub125 μm sieve fraction particles. This suggests that more enzymes can bind to the smaller particles, which is expected since the surface area is higher at a given solid loading. A possible explanation for this difference between the two substrates could be explained by the difference in observed morphologies for each particle preparation. The shredded appearance of the CM-PET likely made an exact measurement of surface area inaccurate, so it is possible that all sieve fractions had much larger surface areas than we estimated. This could explain why the measured invVmax is the same for the CM-PET. Despite this, for both PET particle preparations, these results suggest that decreasing particle size or adding more enzyme can increase the initial reaction rate to a certain extent. It is important to note that this increase in initial rate did not necessarily translate to an increase in overall conversion extent, as mentioned above (Figure 3).

Figure 3

Figure 3. Total amount of monomers released from PET particles hydrolyzed with LCC-ICCG. The bars show the concentrations of all monomers released at the 72 h endpoint of the reaction with indicated [LCC-ICCG] and 10 mg mL–1 PET substrate. Reactions were performed in triplicate, and error bars represent standard deviation. The data shown in this figure are provided in Dataset S3.

Figure 4

Figure 4. InvMM kinetic analysis and total amount of monomers released from PET particles upon hydrolysis with LCC-ICCG. InvMM analysis of 10 mg mL–1 (A) CM-PET and (B) HC-PET hydrolysis rate as a function of LCC-ICCG concentration ([E]). Lines are fits of the invMM equation. Table 2 shows the fitted parameters. Reactions were performed in triplicate and error bars represent standard deviation. The data in this figure are provided in Dataset S5.

Table 2. Kinetic Parameters from InvMM Analysisa
PET substratesieve fraction (μm)invKm (μM)invVmax (μmol g–1 s–1)
CM-PET2503.4 ± 1.70.49 ± 0.14
 1252.7 ± 1.20.48 ± 0.11
 sub1251.1 ± 0.30.40 ± 0.04
HC-PET2500.6 ± 0.10.08 ± 0.01
 1251.3 ± 0.30.14 ± 0.02
 sub1250.7 ± 0.10.16 ± 0.01
a

Calculated from data shown in Figure 3. The error indicates the standard error of the fit.

Since it is known that the optimal pH for LCC-ICCG is ∼8, (17,39) we also evaluated the effect of PET particle size and crystallinity on the performance of LCC-ICCG in bioreactors with pH control. This negates the reduction in enzyme performance due to acidification from the release of the acidic monomers. As reported previously, LCC-ICCG is capable of converting 200 g L–1 amorphized and micronized PET to ∼90% over 10 h when incubated at 72 °C with 3 mg enzyme g PET–1. (39)Figure 5A shows a similar experiment under slightly different conditions, in this case 65 °C and 100 g L–1 PET. A key difference is that the PET was not micronized in this experiment, instead the LC-PET film was cut into ∼1 × 1 cm squares. Using the same enzyme loading of 3 mg g–1 PET, the reaction still reached similar conversion extent in ∼48 h. We also noted a significant lag for the 1 × 1 cm squares which was not observed by Tournier et al. The lag seems to be dependent on surface area and enzyme concentration since a similar lag was observed in the small-scale experiments at low enzyme concentrations and low-surface-area PET particles (Figure S5). While the cause of this induction phase remains unclear, one possible explanation is the enzyme cleaves the PET polymer chains in an internal (endo) manner, as suggested in a previous study investigating the effect of crystallinity on the performance of LCC-ICCG. (65) This would not release monomers in the initial part of the reaction.
There were some interesting differences between the large- and small-scale reactions in the observed MHET concentrations. The large-scale reactions generally built up less MHET, reaching ∼10–12 mM, while the MHET in the small-scale reactions could reach up to ∼21 mM (Figure 2C). This effect was most noticeable with the sub125 μm sieve fraction; however, the large particles in the small-scale reactions generated nearly identical levels of MHET as in the 1 L scale reactions. This suggests that the product released from the PET may be affected by surface area or accessibility of the PET chains. Additionally, since the absolute concentrations of MHET are similar across the large-scale and small-scale experiments with the larger particle sizes, it is possible that at this concentration of MHET, the LCC-ICCG enzyme will preferentially release TPA due to product inhibition. This suggests that MHET is inhibitory and raises the possibility that adding a complementary MHETase to the reaction could further improve the performance of LCC-ICCG, as has been shown with a carboxylesterase and WT-LCC, (66) and the IsPETase and MHETase system. (22,49)

Figure 5

Figure 5. Conversion time courses of 1 L scale reactions conducted in bioreactors. Results from the reaction with various PET substrates at 100 g L–1 and 3 mg g–1 PET LCC-ICCG enzyme. (A–C) Product release profiles as a function of time measured by HPLC for (A) LC-PET film cut into ∼1 × 1 cm squares, (B) HC-PET particles, and (C) WC-PET particles. (D–F) Percent conversion of total PET for (D) LC-PET film cut into ∼1 × 1 cm squares, (E) HC-PET particles, and (F) WC-PET particles. Reactions were performed in duplicate, data points show the mean, and error bars show the absolute difference between the duplicates. The data shown in this figure are provided in Dataset S6.

Conclusions

Click to copy section linkSection link copied!

Overall, under the conditions tested here, we observe that PET particle size does not have an appreciable effect on the overall degradation performance of LCC-ICCG since each particle size fraction for both substrates reached similar extents of degradation. This is in good agreement with previous work from others that investigated the effect of particle size on enzymatic conversion of PET. (52−54) In these studies, the authors generally concluded that smaller PET particles reached higher levels of conversion over days or weeks. However, the enzymes used in these studies, HiC (52,54) and Thc-Cut1, (53) have been reported to exhibit lower hydrolysis rates and reach substantially lower extents of PET conversion compared to LCC-ICCG. (26,57) This suggests that even for reactions spanning days or weeks, these enzymes could still be working at the initial rate. Stated differently, these previous results suggest that across a variety of enzymes working at different rates and extents of conversion, a reduction in particle size may lead only to an increase in the initial rate of the reaction. While this is certainly beneficial to an industrial process since it would lower the residence time in the reactor, based on TEA analysis, this is not as substantial a cost driver as either extent of depolymerization or the cost associated with generating PET feedstocks. (47) For instance, small amorphous particles (<300 μm, as used by Tournier et al. (39)) reach 90% conversion in 10 h. In this study, using the amorphous film cut into small squares, we achieved the same extent of conversion in 48 h (Figure 5). This difference in time would cut the minimum selling price of TPA by less than 6% based on our TEA model. The question then becomes whether the tradeoff in energy consumption (and capital expense) of grinding could be offset by this <6% reduction selling price of TPA─further experiments using LCC-ICCG or other enzymes with various forms of pretreated postconsumer PET waste paired with rigorous TEA will be necessary to determine the efficacy of particle size reduction and amorphization to industrial-scale enzymatic recycling of PET.

Supporting Information

Click to copy section linkSection link copied!

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acssuschemeng.2c01961.

  • Additional characterization and kinetic data as well as datasets for all figures (PDF)

Terms & Conditions

Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.

Author Information

Click to copy section linkSection link copied!

  • Corresponding Authors
    • John E. McGeehan - BOTTLE Consortium, Golden, Colorado 80401, United StatesCentre for Enzyme Innovation, School of Biological Sciences, Institute of Biological and Biomedical Sciences, University of Portsmouth, Portsmouth PO1 2DY, U.K.Orcidhttps://orcid.org/0000-0002-6750-1462 Email: [email protected]
    • Gregg T. Beckham - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United StatesBOTTLE Consortium, Golden, Colorado 80401, United StatesOrcidhttps://orcid.org/0000-0002-3480-212X Email: [email protected]
  • Authors
    • Richard K. Brizendine - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United StatesBOTTLE Consortium, Golden, Colorado 80401, United StatesOrcidhttps://orcid.org/0000-0001-9209-2210
    • Erika Erickson - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United StatesBOTTLE Consortium, Golden, Colorado 80401, United StatesOrcidhttps://orcid.org/0000-0001-7806-9348
    • Stefan J. Haugen - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    • Kelsey J. Ramirez - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United StatesBOTTLE Consortium, Golden, Colorado 80401, United States
    • Joel Miscall - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United StatesBOTTLE Consortium, Golden, Colorado 80401, United States
    • Davinia Salvachúa - Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, Colorado 80401, United States
    • Andrew R. Pickford - BOTTLE Consortium, Golden, Colorado 80401, United StatesCentre for Enzyme Innovation, School of Biological Sciences, Institute of Biological and Biomedical Sciences, University of Portsmouth, Portsmouth PO1 2DY, U.K.Orcidhttps://orcid.org/0000-0002-7237-0030
    • Margaret J. Sobkowicz - Department of Plastics Engineering, University of Massachusetts Lowell, Lowell, Massachusetts 01854, United StatesOrcidhttps://orcid.org/0000-0003-0571-0952
  • Author Contributions

    G.T.B., J.E.M., and M.J.S. conceived the project and R.K.B. and G.T.B. designed the study. R.K.B. and J.M. characterized the PET substrates, R.K.B. and E.E. expressed and purified the enzyme, and R.K.B. conducted the small-scale depolymerization experiments. R.K.B., E.E., and D.S. conducted the bioreactor experiments. S.J.H. and K.J.R. performed all HPLC analyses. R.K.B. and G.T.B. analyzed the data. The paper was written by R.K.B. and G.T.B. and edited and approved by all authors. G.T.B., J.E.M., A.R.P., and M.J.S. were responsible for funding acquisition.

  • Notes
    The authors declare no competing financial interest.

Acknowledgments

Click to copy section linkSection link copied!

Funding was provided by the U.S. DOE, Office of Energy Efficiency and Renewable Energy, Advanced Manufacturing Office (AMO) and Bioenergy Technologies Office (BETO) under contract no. DE-FOA-0002029. This work was also performed as part of the Bio-Optimized Technologies to keep Thermoplastics out of Landfills and the Environment (BOTTLE) Consortium and was supported by AMO and BETO under contract no. DE-AC36-08GO28308 with NREL, operated by Alliance for Sustainable Energy, LLC. The BOTTLE Consortium includes members from the University of Portsmouth, funded under contract no. DE-AC36-08GO28308 with NREL and additionally supported by Research England (E3 scheme). The bioreactor experiments were funded by DARPA via cooperative agreement number IAG-21-17585. This document was approved by DARPA on January 24, 2022, for public release, distribution unlimited. We thank Western Container Corporation for providing PET fines in support of this work. The views expressed in the article do not necessarily represent the views of the DOE or the U.S. Government. The U.S. Government retains and the publisher by accepting the article for publication, acknowledges that the U.S. Government retains a nonexclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this work, or allow others to do so, for U.S. Government purposes.

References

Click to copy section linkSection link copied!

This article references 66 other publications.

  1. 1
    Cózar, A.; Echevarría, F.; González-Gordillo, J. I.; Irigoien, X.; Úbeda, B.; Hernández-León, S.; Palma, Á. T.; Navarro, S.; García-de-Lomas, J.; Ruiz, A. Plastic debris in the open ocean. Proc. Natl. Acad. Sci. U.S.A. 2014, 111, 1023910244,  DOI: 10.1073/pnas.1314705111
  2. 2
    Jambeck, J. R.; Geyer, R.; Wilcox, C.; Siegler, T. R.; Perryman, M.; Andrady, A.; Narayan, R.; Law, K. L. Marine pollution. Plastic waste inputs from land into the ocean. Science 2015, 347, 768771,  DOI: 10.1126/science.1260352
  3. 3
    Allen, S.; Allen, D.; Phoenix, V. R.; Le Roux, G.; Durántez Jiménez, P.; Simonneau, A.; Binet, S.; Galop, D. Atmospheric transport and deposition of microplastics in a remote mountain catchment. Nat. Geosci. 2019, 12, 339344,  DOI: 10.1038/s41561-019-0335-5
  4. 4
    Ellis, L. D.; Rorrer, N. A.; Sullivan, K. P.; Otto, M.; McGeehan, J. E.; Román-Leshkov, Y.; Wierckx, N.; Beckham, G. T. Chemical and biological catalysis for plastics recycling and upcycling. Nat. Catal. 2021, 4, 539556,  DOI: 10.1038/s41929-021-00648-4
  5. 5
    Nicholson, S. R.; Rorrer, N. A.; Carpenter, A. C.; Beckham, G. T. Manufacturing energy and greenhouse gas emissions associated with plastics consumption. Joule 2021, 5, 673686,  DOI: 10.1016/j.joule.2020.12.027
  6. 6
    PET Polymer: Chemical Economics Handbook; IHS Markit. https://ihsmarkit.com/products/pet-polymer-chemical-economics-handbook.html (accessed Sept 01, 2021).
  7. 7
    Ragaert, K.; Delva, L.; Van Geem, K. Mechanical and chemical recycling of solid plastic waste. Waste Manage. 2017, 69, 2458,  DOI: 10.1016/j.wasman.2017.07.044
  8. 8
    Sinha, V.; Patel, M. R.; Patel, J. V. Pet Waste Management by Chemical Recycling: A Review. J. Polym. Environ. 2010, 18, 825,  DOI: 10.1007/s10924-008-0106-7
  9. 9
    Wei, R.; Tiso, T.; Bertling, J.; O’Connor, K.; Blank, L. M.; Bornscheuer, U. T. Possibilities and limitations of biotechnological plastic degradation and recycling. Nat. Catal. 2020, 3, 867871,  DOI: 10.1038/s41929-020-00521-w
  10. 10
    Carniel, A.; Waldow, V. d. A.; Castro, A. M. d. A comprehensive and critical review on key elements to implement enzymatic PET depolymerization for recycling purposes. Biotechnol. Adv. 2021, 52, 107811,  DOI: 10.1016/j.biotechadv.2021.107811
  11. 11
    Wei, R.; Zimmermann, W. Biocatalysis as a green route for recycling the recalcitrant plastic polyethylene terephthalate. Microbiol. Biotechnol. 2017, 10, 13021307,  DOI: 10.1111/1751-7915.12714
  12. 12
    Müller, R.-J.; Schrader, H.; Profe, J.; Dresler, K.; Deckwer, W.-D. Enzymatic Degradation of Poly(ethylene terephthalate): Rapid Hydrolyse using a Hydrolase from T. fusca. Macromol. Rapid Commun. 2005, 26, 14001405,  DOI: 10.1002/marc.200500410
  13. 13
    Alisch-Mark, M.; Herrmann, A.; Zimmermann, W. Increase of the hydrophilicity of polyethylene terephthalate fibres by hydrolases from Thermomonospora fusca and Fusarium solani f. sp. pisi. Biotechnol. Lett. 2006, 28, 681685,  DOI: 10.1007/s10529-006-9041-7
  14. 14
    Eberl, A.; Heumann, S.; Brückner, T.; Araujo, R.; Cavaco-Paulo, A.; Kaufmann, F.; Kroutil, W.; Guebitz, G. M. Enzymatic surface hydrolysis of poly(ethylene terephthalate) and bis(benzoyloxyethyl) terephthalate by lipase and cutinase in the presence of surface active molecules. J. Biotechnol. 2009, 143, 207212,  DOI: 10.1016/j.jbiotec.2009.07.008
  15. 15
    Ronkvist, Å. M.; Xie, W.; Lu, W.; Gross, R. A. Cutinase-Catalyzed Hydrolysis of Poly(ethylene terephthalate). Macromolecules 2009, 42, 51285138,  DOI: 10.1021/ma9005318
  16. 16
    Ribitsch, D.; Heumann, S.; Trotscha, E.; Herrero Acero, E.; Greimel, K.; Leber, R.; Birner-Gruenberger, R.; Deller, S.; Eiteljoerg, I.; Remler, P. Hydrolysis of polyethyleneterephthalate by p-nitrobenzylesterase from Bacillus subtilis. Biotechnol. Prog. 2011, 27, 951960,  DOI: 10.1002/btpr.610
  17. 17
    Sulaiman, S.; Yamato, S.; Kanaya, E.; Kim, J.-J.; Koga, Y.; Takano, K.; Kanaya, S. Isolation of a novel cutinase homolog with polyethylene terephthalate-degrading activity from leaf-branch compost by using a metagenomic approach. Appl. Environ. Microbiol. 2012, 78, 15561562,  DOI: 10.1128/aem.06725-11
  18. 18
    Roth, C.; Wei, R.; Oeser, T.; Then, J.; Föllner, C.; Zimmermann, W.; Sträter, N. Structural and functional studies on a thermostable polyethylene terephthalate degrading hydrolase from Thermobifida fusca. Appl. Microbiol. Biotechnol. 2014, 98, 78157823,  DOI: 10.1007/s00253-014-5672-0
  19. 19
    Sulaiman, S.; You, D.-J.; Kanaya, E.; Koga, Y.; Kanaya, S. Crystal Structure and Thermodynamic and Kinetic Stability of Metagenome-Derived LC-Cutinase. Biochemistry 2014, 53, 18581869,  DOI: 10.1021/bi401561p
  20. 20
    Wei, R.; Oeser, T.; Zimmermann, W. Synthetic polyester-hydrolyzing enzymes from thermophilic actinomycetes. Adv. Appl. Microbiol. 2014, 89, 267305,  DOI: 10.1016/B978-0-12-800259-9.00007-X
  21. 21
    Perz, V.; Bleymaier, K.; Sinkel, C.; Kueper, U.; Bonnekessel, M.; Ribitsch, D.; Guebitz, G. M. Substrate specificities of cutinases on aliphatic-aromatic polyesters and on their model substrates. New Biotechnol. 2016, 33, 295304,  DOI: 10.1016/j.nbt.2015.11.004
  22. 22
    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, 11961199,  DOI: 10.1126/science.aad6359
  23. 23
    Ribitsch, D.; Hromic, A.; Zitzenbacher, S.; Zartl, B.; Gamerith, C.; Pellis, A.; Jungbauer, A.; Łyskowski, A.; Steinkellner, G.; Gruber, K. Small cause, large effect: Structural characterization of cutinases from Thermobifida cellulosilytica. Biotechnol. Bioeng. 2017, 114, 24812488,  DOI: 10.1002/bit.26372
  24. 24
    Kawai, F.; Kawabata, T.; Oda, M. Current State and Perspectives Related to the Polyethylene Terephthalate Hydrolases Available for Biorecycling. ACS Sustainable Chem. Eng. 2020, 8, 88948908,  DOI: 10.1021/acssuschemeng.0c01638
  25. 25
    Sonnendecker, C.; Oeser, J.; Richter, P. K.; Hille, P.; Zhao, Z.; Fischer, C.; Lippold, H.; Blazquez-Sanchez, P.; Engelberger, F.; Ramirez-Sarmiento, C. A. Low Carbon Footprint Recycling of Post-Consumer PET Plastic with a Metagenomic Polyester Hydrolase. ChemSusChem 2022, 15, e202101062  DOI: 10.1002/cssc.202101062
  26. 26
    Herrero Acero, E.; Ribitsch, D.; Dellacher, A.; Zitzenbacher, S.; Marold, A.; Steinkellner, G.; Gruber, K.; Schwab, H.; Guebitz, G. M. Surface engineering of a cutinase from Thermobifida cellulosilytica for improved polyester hydrolysis. Biotechnol. Bioeng. 2013, 110, 25812590,  DOI: 10.1002/bit.24930
  27. 27
    Ribitsch, D.; Yebra, A. O.; Zitzenbacher, S.; Wu, J.; Nowitsch, S.; Steinkellner, G.; Greimel, K.; Doliska, A.; Oberdorfer, G.; Gruber, C. C. Fusion of binding domains to Thermobifida cellulosilytica cutinase to tune sorption characteristics and enhancing PET hydrolysis. Biomacromolecules 2013, 14, 17691776,  DOI: 10.1021/bm400140u
  28. 28
    Ribitsch, D.; Herrero Acero, E.; Przylucka, A.; Zitzenbacher, S.; Marold, A.; Gamerith, C.; Tscheliessnig, R.; Jungbauer, A.; Rennhofer, H.; Lichtenegger, H. Enhanced cutinase-catalyzed hydrolysis of polyethylene terephthalate by covalent fusion to hydrophobins. Appl. Environ. Microbiol. 2015, 81, 35863592,  DOI: 10.1128/AEM.04111-14
  29. 29
    Then, J.; Wei, R.; Oeser, T.; Barth, M.; Belisário-Ferrari, M. R.; Schmidt, J.; Zimmermann, W. Ca2+ and Mg2+ binding site engineering increases the degradation of polyethylene terephthalate films by polyester hydrolases from Thermobifida fusca. Biotechnol. J. 2015, 10, 592598,  DOI: 10.1002/biot.201400620
  30. 30
    Wei, R.; Oeser, T.; Schmidt, J.; Meier, R.; Barth, M.; Then, J.; Zimmermann, W. Engineered bacterial polyester hydrolases efficiently degrade polyethylene terephthalate due to relieved product inhibition. Biotechnol. Bioeng. 2016, 113, 16581665,  DOI: 10.1002/bit.25941
  31. 31
    Shirke, A. N.; Basore, D.; Butterfoss, G. L.; Bonneau, R.; Bystroff, C.; Gross, R. A. Toward rational thermostabilization of Aspergillus oryzae cutinase: Insights into catalytic and structural stability. Proteins 2016, 84, 6072,  DOI: 10.1002/prot.24955
  32. 32
    Biundo, A.; Ribitsch, D.; Steinkellner, G.; Gruber, K.; Guebitz, G. M. Polyester hydrolysis is enhanced by a truncated esterase: Less is more. Biotechnol. J. 2017, 12,  DOI: 10.1002/biot.201600450
  33. 33
    Shirke, A. N.; Butterfoss, G. L.; Saikia, R.; Basu, A.; Maria, L.; Svendsen, A.; Gross, R. A. Engineered Humicola insolens cutinase for efficient cellulose acetate deacetylation. Biotechnol. J. 2017, 12, 1700188,  DOI: 10.1002/biot.201700188
  34. 34
    Austin, H. P.; Allen, M. D.; Donohoe, B. S.; Rorrer, N. A.; Kearns, F. L.; Silveira, R. L.; Pollard, B. C.; Dominick, G.; Duman, R.; El Omari, K. Characterization and engineering of a plastic-degrading aromatic polyesterase. Proc. Natl. Acad. Sci. U.S.A. 2018, 115, E4350E4357,  DOI: 10.1073/pnas.1718804115
  35. 35
    Biundo, A.; Ribitsch, D.; Guebitz, G. M. Surface engineering of polyester-degrading enzymes to improve efficiency and tune specificity. Appl. Microbiol. Biotechnol. 2018, 102, 35513559,  DOI: 10.1007/s00253-018-8850-7
  36. 36
    Shirke, A. N.; White, C.; Englaender, J. A.; Zwarycz, A.; Butterfoss, G. L.; Linhardt, R. J.; Gross, R. A. Stabilizing Leaf and Branch Compost Cutinase (LCC) with Glycosylation: Mechanism and Effect on PET Hydrolysis. Biochemistry 2018, 57, 11901200,  DOI: 10.1021/acs.biochem.7b01189
  37. 37
    Son, H. F.; Cho, I. J.; Joo, S.; Seo, H.; Sagong, H.-Y.; Choi, S. Y.; Lee, S. Y.; Kim, K.-J. Rational Protein Engineering of Thermo-Stable PETase from Ideonella sakaiensis for Highly Efficient PET Degradation. ACS Catal. 2019, 9, 35193526,  DOI: 10.1021/acscatal.9b00568
  38. 38
    Furukawa, M.; Kawakami, N.; Tomizawa, A.; Miyamoto, K. Efficient Degradation of Poly(ethylene terephthalate) with Thermobifida fusca Cutinase Exhibiting Improved Catalytic Activity Generated using Mutagenesis and Additive-based Approaches. Sci. Rep. 2019, 9, 16038,  DOI: 10.1038/s41598-019-52379-z
  39. 39
    Tournier, V.; Topham, C. M.; Gilles, A.; David, B.; Folgoas, C.; Moya-Leclair, E.; Kamionka, E.; Desrousseaux, M.-L.; Texier, H.; Gavalda, S. An engineered PET depolymerase to break down and recycle plastic bottles. Nature 2020, 580, 216219,  DOI: 10.1038/s41586-020-2149-4
  40. 40
    Bell, E.; Smithson, R.; Kilbride, S.; Foster, J.; Hardy, F.; Ramachandran, S.; Tedstone, A.; Haigh, S.; Garforth, A.; Day, P. Directed Evolution of an Efficient and Thermostable PET Depolymerase. ChemRxiv 2021,  DOI: 10.26434/chemrxiv-2021-mcjh6
  41. 41
    Cui, Y.; Chen, Y.; Liu, X.; Dong, S.; Tian, Y. e.; Qiao, Y.; Mitra, R.; Han, J.; Li, C.; Han, X. Computational Redesign of a PETase for Plastic Biodegradation under Ambient Condition by the GRAPE Strategy. ACS Catal. 2021, 11, 13401350,  DOI: 10.1021/acscatal.0c05126
  42. 42
    Nakamura, A.; Kobayashi, N.; Koga, N.; Iino, R. Positive Charge Introduction on the Surface of Thermostabilized PET Hydrolase Facilitates PET Binding and Degradation. ACS Catal. 2021, 11, 85508564,  DOI: 10.1021/acscatal.1c01204
  43. 43
    Guo, B.; Vanga, S. R.; Lopez-Lorenzo, X.; Saenz-Mendez, P.; Ericsson, S. R.; Fang, Y.; Ye, X.; Schriever, K.; Bäckström, E.; Biundo, A. Conformational Selection in Biocatalytic Plastic Degradation by PETase. ACS Catal. 2022, 12, 33973409,  DOI: 10.1021/acscatal.1c05548
  44. 44
    Lu, H.; Diaz, D. J.; Czarnecki, N. J.; Zhu, C.; Kim, W.; Shroff, R.; Acosta, D. J.; Alexander, B. R.; Cole, H. O.; Zhang, Y. Machine learning-aided engineering of hydrolases for PET depolymerization. Nature 2022, 604, 662667,  DOI: 10.1038/s41586-022-04599-z
  45. 45
    Wei, R.; von Haugwitz, G.; Pfaff, L.; Mican, J.; Badenhorst, C. P. S.; Liu, W.; Weber, G.; Austin, H. P.; Bednar, D.; Damborsky, J. Mechanism-Based Design of Efficient PET Hydrolases. ACS Catal. 2022, 12, 33823396,  DOI: 10.1021/acscatal.1c05856
  46. 46
    Zeng, W.; Li, X.; Yang, Y.; Min, J.; Huang, J.-W.; Liu, W.; Niu, D.; Yang, X.; Han, X.; Zhang, L. Substrate-Binding Mode of a Thermophilic PET Hydrolase and Engineering the Enzyme to Enhance the Hydrolytic Efficacy. ACS Catal. 2022, 12, 30333040,  DOI: 10.1021/acscatal.1c05800
  47. 47
    Singh, A.; Rorrer, N. A.; Nicholson, S. R.; Erickson, E.; DesVeaux, J. S.; Avelino, A. F. T.; Lamers, P.; Bhatt, A.; Zhang, Y.; Avery, G. Techno-economic, life-cycle, and socioeconomic impact analysis of enzymatic recycling of poly(ethylene terephthalate). Joule 2021, 5, 24792503,  DOI: 10.1016/j.joule.2021.06.015
  48. 48
    Wei, R.; Breite, D.; Song, C.; Gräsing, D.; Ploss, T.; Hille, P.; Schwerdtfeger, R.; Matysik, J.; Schulze, A.; Zimmermann, W. Biocatalytic Degradation Efficiency of Postconsumer Polyethylene Terephthalate Packaging Determined by Their Polymer Microstructures. Adv. Sci. 2019, 6, 1900491,  DOI: 10.1002/advs.201900491
  49. 49
    Knott, B. C.; Erickson, E.; Allen, M. D.; Gado, J. E.; Graham, R.; Kearns, F. L.; Pardo, I.; Topuzlu, E.; Anderson, J. J.; Austin, H. P. Characterization and engineering of a two-enzyme system for plastics depolymerization. Proc. Natl. Acad. Sci. U.S.A. 2020, 117, 2547625485,  DOI: 10.1073/pnas.2006753117
  50. 50
    Erickson, E.; Shakespeare, T. J.; Bratti, F.; Buss, B. L.; Graham, R.; Hawkins, M. A.; König, G.; Michener, W. E.; Miscall, J.; Ramirez, K. J. Comparative performance of PETase as a function of reaction conditions, substrate properties, and product accumulation. ChemSusChem 2022, 15, e202101932  DOI: 10.1002/cssc.202101932
  51. 51
    Lu, H.; Diaz, D. J.; Czarnecki, N. J.; Zhu, C.; Kim, W.; Shroff, R.; Acosta, D. J.; Alexander, B.; Cole, H.; Zhang, Y. J. Deep learning redesign of PETase for practical PET degrading applications. bioRxiv 2021,  DOI: 10.1101/2021.10.10.463845
  52. 52
    de Castro, A. M.; Carniel, A.; Nicomedes Junior, J.; da Conceição Gomes, A.; Valoni, É. Screening of commercial enzymes for poly(ethylene terephthalate) (PET) hydrolysis and synergy studies on different substrate sources. J. Ind. Microbiol. Biotechnol. 2017, 44, 835844,  DOI: 10.1007/s10295-017-1942-z
  53. 53
    Gamerith, C.; Zartl, B.; Pellis, A.; Guillamot, F.; Marty, A.; Acero, E. H.; Guebitz, G. M. Enzymatic recovery of polyester building blocks from polymer blends. Process Biochem. 2017, 59, 5864,  DOI: 10.1016/j.procbio.2017.01.004
  54. 54
    Castro, A. M. d.; Carniel, A.; Stahelin, D.; Chinelatto Junior, L. S.; Honorato, H. d. A.; de Menezes, S. M. C. High-fold improvement of assorted post-consumer poly(ethylene terephthalate) (PET) packages hydrolysis using Humicola insolens cutinase as a single biocatalyst. Process Biochem. 2019, 81, 8591,  DOI: 10.1016/j.procbio.2019.03.006
  55. 55
    Kari, J.; Andersen, M.; Borch, K.; Westh, P. An Inverse Michaelis–Menten Approach for Interfacial Enzyme Kinetics. ACS Catal. 2017, 7, 49044914,  DOI: 10.1021/acscatal.7b00838
  56. 56
    Andersen, M.; Kari, J.; Borch, K.; Westh, P. Michaelis-Menten equation for degradation of insoluble substrate. Math. Biosci. 2018, 296, 9397,  DOI: 10.1016/j.mbs.2017.11.011
  57. 57
    Bååth, J. A.; Borch, K.; Jensen, K.; Brask, J.; Westh, P. Comparative Biochemistry of Four Polyester (PET) Hydrolases. ChemBioChem 2021, 22, 16271637,  DOI: 10.1002/cbic.202000793
  58. 58
    Werner, A. Z.; Clare, R.; Mand, T. D.; Pardo, I.; Ramirez, K. J.; Haugen, S. J.; Bratti, F.; Dexter, G. N.; Elmore, J. R.; Huenemann, J. D. Tandem chemical deconstruction and biological upcycling of poly(ethylene terephthalate) to beta-ketoadipic acid by Pseudomonas putida KT2440. Metab. Eng. 2021, 67, 250261,  DOI: 10.1016/j.ymben.2021.07.005
  59. 59
    Blundell, D. J.; MacKerron, D. H.; Fuller, W.; Mahendrasingam, A.; Martin, C.; Oldman, R. J.; Rule, R. J.; Riekel, C. Characterization of strain-induced crystallization of poly(ethylene terephthalate) at fast draw rates using synchrotron radiation. Polymer 1996, 37, 33033311,  DOI: 10.1016/0032-3861(96)88476-X
  60. 60
    Blundell, D. J.; Mahendrasingam, A.; Martin, C.; Fuller, W.; MacKerron, D. H.; Harvie, J. L.; Oldman, R. J.; Riekel, C. Orientation prior to crystallisation during drawing of poly(ethylene terephthalate). Polymer 2000, 41, 77937802,  DOI: 10.1016/S0032-3861(00)00128-2
  61. 61
    Mahendrasingam, A.; Martin, C.; Fuller, W.; Blundell, D. J.; Oldman, R. J.; MacKerron, D. H.; Harvie, J. L.; Riekel, C. Observation of a transient structure prior to strain-induced crystallization in poly(ethylene terephthalate). Polymer 2000, 41, 12171221,  DOI: 10.1016/S0032-3861(99)00461-9
  62. 62
    Forestier, E.; Combeaud, C.; Guigo, N.; Sbirrazzuoli, N.; Billon, N. Understanding of strain-induced crystallization developments scenarios for polyesters: Comparison of poly(ethylene furanoate), PEF, and poly(ethylene terephthalate), PET. Polymer 2020, 203, 122755,  DOI: 10.1016/j.polymer.2020.122755
  63. 63
    Bashir, Z.; Al-Aloush, I.; Al-Raqibah, I.; Ibrahim, M. Evaluation of three methods for the measurement of crystallinity of PET resins, preforms, and bottles. Polym. Eng. Sci. 2000, 40, 24422455,  DOI: 10.1002/pen.11376
  64. 64
    Scandola, M.; Focarete, M. L.; Frisoni, G. Simple Kinetic Model for the Heterogeneous Enzymatic Hydrolysis of Natural Poly(3-hydroxybutyrate). Macromolecules 1998, 31, 38463851,  DOI: 10.1021/ma980137y
  65. 65
    Thomsen, T. B.; Hunt, C. J.; Meyer, A. S. Influence of substrate crystallinity and glass transition temperature on enzymatic degradation of polyethylene terephthalate (PET). New Biotechnol. 2022, 69, 2835,  DOI: 10.1016/j.nbt.2022.02.006
  66. 66
    Barth, M.; Honak, A.; Oeser, T.; Wei, R.; Belisário-Ferrari, M. R.; Then, J.; Schmidt, J.; Zimmermann, W. A dual enzyme system composed of a polyester hydrolase and a carboxylesterase enhances the biocatalytic degradation of polyethylene terephthalate films. Biotechnol. J. 2016, 11, 10821087,  DOI: 10.1002/biot.201600008

Cited By

Click to copy section linkSection link copied!
Citation Statements
Explore this article's citation statements on scite.ai

This article is cited by 62 publications.

  1. Daria Lazarenko, Graham P. Schmidt, Michael F. Crowley, Gregg T. Beckham, Brandon C. Knott. Molecular Details of Polyester Decrystallization via Molecular Simulation. Macromolecules 2025, Article ASAP.
  2. Suprio Kamal, Kiersten Carew, Ji Qin, Dilara Hatinoglu, Onur Apul, Boya Xiong. Combined Adsorption and Michaelis–Menten Approach Reveals Predominant Enzymatic Depolymerization of Crystalline Poly(ethylene terephthalate) by Humicola insolens Cutinase in the Solution Phase. ACS Sustainable Chemistry & Engineering 2025, Article ASAP.
  3. Hwidong Hwang, Myungwan Han. Kinetics of Polyester Methanolysis by a Consecutive Reaction Model. Industrial & Engineering Chemistry Research 2025, 64 (2) , 1085-1094. https://doi.org/10.1021/acs.iecr.4c03791
  4. Thomas M. Groseclose, Erin A. Kober, Matilda Clark, Benjamin Moore, Shounak Banerjee, Victoria Bemmer, Gregg T. Beckham, Andrew R. Pickford, Taraka T. Dale, Hau B. Nguyen. A High-Throughput Screening Platform for Engineering Poly(ethylene Terephthalate) Hydrolases. ACS Catalysis 2024, 14 (19) , 14622-14638. https://doi.org/10.1021/acscatal.4c04321
  5. Mackenzie C. R. Denton, Natasha P. Murphy, Brenna Norton-Baker, Mauro Lua, Harrison Steel, Gregg T. Beckham. Integration of pH Control into Chi.Bio Reactors and Demonstration with Small-Scale Enzymatic Poly(ethylene terephthalate) Hydrolysis. Biochemistry 2024, 63 (13) , 1599-1607. https://doi.org/10.1021/acs.biochem.4c00149
  6. Virender Kumar, Alessandro Pellis, Reinhard Wimmer, Vladimir Popok, Jesper de Claville Christiansen, Cristiano Varrone. Efficient Depolymerization of Poly(ethylene 2,5-furanoate) Using Polyester Hydrolases. ACS Sustainable Chemistry & Engineering 2024, 12 (26) , 9658-9668. https://doi.org/10.1021/acssuschemeng.4c00915
  7. Lakshmiprasad Gurrala, Rafi Anowar, Ana Rita C. Morais. Probing Supercritical CO2 to Improve the Susceptibility of Semicrystalline Polyethylene Terephthalate to Enzymatic Hydrolysis. ACS Sustainable Chemistry & Engineering 2024, 12 (20) , 7713-7723. https://doi.org/10.1021/acssuschemeng.3c08286
  8. Adam McNeeley, Y. A. Liu. Assessment of PET Depolymerization Processes for Circular Economy. 1. Thermodynamics, Chemistry, Purification, and Process Design. Industrial & Engineering Chemistry Research 2024, 63 (8) , 3355-3399. https://doi.org/10.1021/acs.iecr.3c04000
  9. Mario A. Cribari, Maxwell J. Unger, Ilona C. Unarta, Ashley N. Ogorek, Xuhui Huang, Jeffrey D. Martell. Ultrahigh-Throughput Directed Evolution of Polymer-Degrading Enzymes Using Yeast Display. Journal of the American Chemical Society 2023, 145 (50) , 27380-27389. https://doi.org/10.1021/jacs.3c08291
  10. Grégory Arnal, Julien Anglade, Sabine Gavalda, Vincent Tournier, Nicolas Chabot, Uwe T. Bornscheuer, Gert Weber, Alain Marty. Assessment of Four Engineered PET Degrading Enzymes Considering Large-Scale Industrial Applications. ACS Catalysis 2023, 13 (20) , 13156-13166. https://doi.org/10.1021/acscatal.3c02922
  11. Vincent Tournier, Sophie Duquesne, Frédérique Guillamot, Henri Cramail, Daniel Taton, Alain Marty, Isabelle André. Enzymes’ Power for Plastics Degradation. Chemical Reviews 2023, 123 (9) , 5612-5701. https://doi.org/10.1021/acs.chemrev.2c00644
  12. Akanksha Patel, Allen C. Chang, Sarah Perry, Ya-Hue V. Soong, Christian Ayafor, Hsi-Wu Wong, Dongming Xie, Margaret J. Sobkowicz. Melt Processing Pretreatment Effects on Enzymatic Depolymerization of Poly(ethylene terephthalate). ACS Sustainable Chemistry & Engineering 2022, 10 (41) , 13619-13628. https://doi.org/10.1021/acssuschemeng.2c03142
  13. Ya-Jun Liu, Fei Yan, Weiliang Dong, Yuman Sun, Ren Wei, Yingang Feng. Optimized whole-cell depolymerization of polyethylene terephthalate to monomers using engineered Clostridium thermocellum. Journal of Hazardous Materials 2025, 488 , 137441. https://doi.org/10.1016/j.jhazmat.2025.137441
  14. Shaobin Zhang, Xuan Wang, Haixia Shen, Jing Zhang, Weiliang Dong, Ziyi Yu. Scalable nanoplastic degradation in water with enzyme-functionalized porous hydrogels. Journal of Hazardous Materials 2025, 487 , 137196. https://doi.org/10.1016/j.jhazmat.2025.137196
  15. Ali Zaker, Karine Auclair. Impact of Ball Milling on the Microstructure of Polyethylene Terephthalate. ChemSusChem 2025, 18 (4) https://doi.org/10.1002/cssc.202401506
  16. Christian Ayafor, Allen C. Chang, Akanksha Patel, Umer Abid, Dongming Xie, Margaret J. Sobkowicz, Hsi‐Wu Wong. In‐Situ Product Removal for the Enzymatic Depolymerization of Poly(ethylene terephthalate) via a Membrane Reactor. ChemSusChem 2025, 18 (3) https://doi.org/10.1002/cssc.202400698
  17. Yongjie Wang, Ekram Akram, Yujing Ding, Chengzhi He, Yifei Zhang. Fusion of Hydrophobic Anchor Peptides Promotes the Hydrolytic Activity of PETase but not the Extent of PET Depolymerization. ChemCatChem 2025, 17 (3) https://doi.org/10.1002/cctc.202401252
  18. Zdeněk Petrášek, Bernd Nidetzky. The Effect of Accessibility of Insoluble Substrate on the Overall Kinetics of Enzymatic Degradation. Biotechnology and Bioengineering 2025, 7 https://doi.org/10.1002/bit.28921
  19. Yu Zhou, Jinxu Zhang, Yunxin Zheng, Wei Lin, Shengping You, Mengfan Wang, Rongxin Su, Wei Qi. Simple enzymatic depolymerization process based on rapid ball milling pretreatment for high-crystalline polyethylene terephthalate fibers. Bioresource Technology 2025, 416 , 131759. https://doi.org/10.1016/j.biortech.2024.131759
  20. Tobias Heinks, Katrin Hofmann, Lennard Zimmermann, Igor Gamm, Alexandra Lieb, Luise Blach, Ren Wei, Uwe T. Bornscheuer, Julian Thiele, Christof Hamel, Jan von Langermann. Analysis of the product-spectrum during the biocatalytic hydrolysis of PEF (poly(ethylene furanoate)) with various esterases. RSC Sustainability 2025, 123 https://doi.org/10.1039/D4SU00722K
  21. Sacha Pérocheau Arnaud, Véronique Michelet, Sandra Olivero, Patrick Navard, Christelle Combeaud, Alice Mija. New method for extraction, identification, and quantification of non‐intentionally added substances in polystyrene. Journal of Vinyl and Additive Technology 2024, https://doi.org/10.1002/vnl.22188
  22. Yu Zhou, Jiaxing Zhang, Shengping You, Wei Lin, Baoyu Zhang, Mengfan Wang, Rongxin Su, Wei Qi. High terephthalic acid purity: Effective polyethylene terephthalate degradation process based on pH regulation with dual-function hydrolase. Bioresource Technology 2024, 413 , 131461. https://doi.org/10.1016/j.biortech.2024.131461
  23. Stefanie Fritzsche, Holger Hübner, Marco Oldiges, Kathrin Castiglione. Comparative evaluation of the extracellular production of a polyethylene terephthalate degrading cutinase by Corynebacterium glutamicum and leaky Escherichia coli in batch and fed-batch processes. Microbial Cell Factories 2024, 23 (1) https://doi.org/10.1186/s12934-024-02547-2
  24. Wei Liu, Chuang Li, Bin Li, Liying Zhu, Dengming Ming, Ling Jiang. Structure-guided discovery and rational design of a new poly(ethylene terephthalate) hydrolase from AlphaFold protein structure database. Journal of Hazardous Materials 2024, 480 , 136389. https://doi.org/10.1016/j.jhazmat.2024.136389
  25. Matthew Colachis, Jacob L. Lilly, Edward Trigg, Katarzyna H. Kucharzyk. Analytical tools to assess polymer biodegradation: A critical review and recommendations. Science of The Total Environment 2024, 955 , 176920. https://doi.org/10.1016/j.scitotenv.2024.176920
  26. Akanksha Patel, Allen C. Chang, Umer Abid, Christian Ayafor, Hsi‐Wu Wong, Dongming Xie, Margaret J. Sobkowicz. Enzymatic depolymerization of polyester: Foaming as a pretreatment to increase specific surface area. Journal of Applied Polymer Science 2024, 141 (44) https://doi.org/10.1002/app.56177
  27. Ximena Lopez-Lorenzo, David Hueting, Eliott Bosshard, Per-Olof Syrén. Degradation of PET microplastic particles to monomers in human serum by PETase. Faraday Discussions 2024, 252 , 387-402. https://doi.org/10.1039/D4FD00014E
  28. Yuhong Cheng, Yihao Cheng, Shengcheng Zhou, Yelizhati Ruzha, Yu Yang. Closed-loop recycling of PET fabric and bottle waste by tandem pre-amorphization and enzymatic hydrolysis. Resources, Conservation and Recycling 2024, 208 , 107706. https://doi.org/10.1016/j.resconrec.2024.107706
  29. Hendrik Puetz, Alexander-Maurice Illig, Mariia Vorobii, Christoph Janknecht, Francisca Contreras, Fabian Flemig, Ulrich Schwaneberg. KnowVolution of an efficient polyamidase through molecular dynamics simulations of incrementally docked oligomeric substrates. 2024https://doi.org/10.1101/2024.08.13.607760
  30. Thi Kim Anh Nguyen, Thành Trần‐Phú, Rahman Daiyan, Xuan Minh Chau Ta, Rose Amal, Antonio Tricoli. From Plastic Waste to Green Hydrogen and Valuable Chemicals Using Sunlight and Water. Angewandte Chemie 2024, 136 (32) https://doi.org/10.1002/ange.202401746
  31. Thi Kim Anh Nguyen, Thành Trần‐Phú, Rahman Daiyan, Xuan Minh Chau Ta, Rose Amal, Antonio Tricoli. From Plastic Waste to Green Hydrogen and Valuable Chemicals Using Sunlight and Water. Angewandte Chemie International Edition 2024, 63 (32) https://doi.org/10.1002/anie.202401746
  32. Akanksha Patel, Allen C. Chang, Umer Abid, Christian Ayafor, Hsi-Wu Wong, Dongming Xie, Margaret J. Sobkowicz. Effects of Copolymer Structure on Enzyme-Catalyzed Polyester Recycling. Journal of Polymers and the Environment 2024, 32 (8) , 3961-3972. https://doi.org/10.1007/s10924-024-03223-7
  33. Matthew Colachis, Nathan Clark, Ashley Frank, Edward B. Trigg, Colin Hinton, Greg Gregoriades, Vance Gustin, Ryan Daly, Rachel Thurston, Bryon Moore, Katarzyna H. Kucharzyk, Jacob L. Lilly. An efficient and scalable melt fiber spinning system to improve enzyme-based PET recycling. Chemical Engineering Journal Advances 2024, 19 , 100624. https://doi.org/10.1016/j.ceja.2024.100624
  34. Chengyong Wang, Rui Long, Xiran Lin, Wei Liu, Liying Zhu, Ling Jiang. Development and characterization of a bacterial enzyme cascade reaction system for efficient and stable PET degradation. Journal of Hazardous Materials 2024, 472 , 134480. https://doi.org/10.1016/j.jhazmat.2024.134480
  35. Amy A. Cuthbertson, Clarissa Lincoln, Joel Miscall, Lisa M. Stanley, Anjani K. Maurya, Arun S. Asundi, Christopher J. Tassone, Nicholas A. Rorrer, Gregg T. Beckham. Characterization of polymer properties and identification of additives in commercially available research plastics. Green Chemistry 2024, 26 (12) , 7067-7090. https://doi.org/10.1039/D4GC00659C
  36. Yu-Ji Luo, Jia-Yin Sun, Zhi Li. Rapid chemical recycling of waste polyester plastics catalyzed by recyclable catalyst. Green Chemical Engineering 2024, 5 (2) , 257-265. https://doi.org/10.1016/j.gce.2023.06.002
  37. Sune W. Schubert, Thore B. Thomsen, Kristine S. Clausen, Anders Malmendal, Cameron J. Hunt, Kim Borch, Kenneth Jensen, Jesper Brask, Anne S. Meyer, Peter Westh. Relationships of crystallinity and reaction rates for enzymatic degradation of poly (ethylene terephthalate), PET. ChemSusChem 2024, 17 (10) https://doi.org/10.1002/cssc.202301752
  38. Yvonne Joho, Santana Royan, Alessandro T. Caputo, Sophia Newton, Thomas S. Peat, Janet Newman, Colin Jackson, Albert Ardevol. Enhancing PET Degrading Enzymes: A Combinatory Approach. ChemBioChem 2024, 25 (10) https://doi.org/10.1002/cbic.202400084
  39. Camila Guajardo, Rodrigo Andler. Challenges and perspectives in enzymatic polymer fragmentation: The case of rubber and polyethylene terephthalate. Journal of Cleaner Production 2024, 450 , 141875. https://doi.org/10.1016/j.jclepro.2024.141875
  40. Ximena Lopez-Lorenzo, David Hueting, Eliott Bosshard, Per-Olof Syrén. WITHDRAWN: Degradation of PET microplastic particles to monomers in human serum by PETase. 2024https://doi.org/10.21203/rs.3.rs-3164368/v2
  41. Mackenzie C.R. Denton, Natasha P. Murphy, Brenna Norton-Baker, Mauro Lua, Harrison Steel, Gregg T. Beckham. Integration of pH control into Chi.Bio reactors and demonstration with small-scale enzymatic poly(ethylene terephthalate) hydrolysis. 2024https://doi.org/10.1101/2024.03.03.582641
  42. Lei He, Shan-Shan Yang, Jie Ding, Cheng-Xin Chen, Fan Yang, Zhi-Li He, Ji-Wei Pang, Bo-Yu Peng, Yalei Zhang, De-Feng Xing, Nan-Qi Ren, Wei-Min Wu. Biodegradation of polyethylene terephthalate by Tenebrio molitor: Insights for polymer chain size, gut metabolome and host genes. Journal of Hazardous Materials 2024, 465 , 133446. https://doi.org/10.1016/j.jhazmat.2024.133446
  43. Thore Bach Thomsen, Tobias S. Radmer, Anne S. Meyer. Enzymatic degradation of poly(ethylene terephthalate) (PET): Identifying the cause of the hypersensitive enzyme kinetic response to increased PET crystallinity. Enzyme and Microbial Technology 2024, 173 , 110353. https://doi.org/10.1016/j.enzmictec.2023.110353
  44. Lixia Shi, Leilei Zhu. Recent Advances and Challenges in Enzymatic Depolymerization and Recycling of PET Wastes. ChemBioChem 2024, 25 (2) https://doi.org/10.1002/cbic.202300578
  45. Chengyong Wang, Rui Long, Xiran Lin, Wei Liu, Liying Zhu, Ling Jiang. Development and Characterization of a Bacterial Enzyme Cascade Reaction System for Efficient and Stable Pet Degradation. 2024https://doi.org/10.2139/ssrn.4755922
  46. Dong Lu, Jinglong Wu, Shuming Jin, Qiuyang Wu, Li Deng, Fang Wang, Kaili Nie. The enhancement of waste PET particles enzymatic degradation with a rotating packed bed reactor. Journal of Cleaner Production 2024, 434 , 140088. https://doi.org/10.1016/j.jclepro.2023.140088
  47. Ya‐Hue Valerie Soong, Umer Abid, Allen C. Chang, Christian Ayafor, Akanksha Patel, Jiansong Qin, Jin Xu, Carl Lawton, Hsi‐Wu Wong, Margaret J. Sobkowicz, Dongming Xie. Enzyme selection, optimization, and production toward biodegradation of post‐consumer poly(ethylene terephthalate) at scale. Biotechnology Journal 2023, 18 (12) https://doi.org/10.1002/biot.202300119
  48. Siwen Bi, Zhuang Zhang, Zhenzhen Yang, Zitong Shen, Jiahui Cai, Jintao Hu, Haoxiang Jin, Tianhao Qiu, Peng Yu, Bin Tan. Protein modified cellulose nanocrystals on reinforcement and self-driven biodegradation of aliphatic polyester. Carbohydrate Polymers 2023, 322 , 121312. https://doi.org/10.1016/j.carbpol.2023.121312
  49. Thore Bach Thomsen, Kristoffer Almdal, Anne S. Meyer. Significance of poly(ethylene terephthalate) (PET) substrate crystallinity on enzymatic degradation. New Biotechnology 2023, 78 , 162-172. https://doi.org/10.1016/j.nbt.2023.11.001
  50. Jun Zhang, Hongzhao Wang, Zhaorong Luo, Zhenwu Yang, Zixuan Zhang, Pengyu Wang, Mengyu Li, Yi Zhang, Yue Feng, Diannan Lu, Yushan Zhu. Computational design of highly efficient thermostable MHET hydrolases and dual enzyme system for PET recycling. Communications Biology 2023, 6 (1) https://doi.org/10.1038/s42003-023-05523-5
  51. Sune W. Schubert, Thore B. Thomsen, Kristine S. Clausen, Anders Malmendal, Cameron J. Hunt, Kim Borch, Kenneth Jensen, Jesper Brask, Anne S. Meyer, Peter Westh. Relationships of crystallinity and reaction rates for enzymatic degradation of poly (ethylene terephthalate), PET. 2023https://doi.org/10.1101/2023.11.05.564366
  52. Shan-Shan Yang, Wei-Min Wu, Ji-Wei Pang, Lei He, Meng-Qi Ding, Mei-Xi Li, Yi-Lin Zhao, Han-Jun Sun, De-Feng Xing, Nan-Qi Ren, Jun Yang, Craig S. Criddle, Jie Ding. Bibliometric analysis of publications on biodegradation of plastics: Explosively emerging research over 70 years. Journal of Cleaner Production 2023, 428 , 139423. https://doi.org/10.1016/j.jclepro.2023.139423
  53. Akanksha Patel, Allen C. Chang, Abigail Mastromonaco, Mauricio Acosta Diaz, Sarah Perry, Olivia Ferki, Christian Ayafor, Umer Abid, Hsi-Wu Wong, Dongming Xie, Margaret J. Sobkowicz. Aqueous buffer solution-induced crystallization competes with enzymatic depolymerization of pre-treated post-consumer poly (ethylene terephthalate) waste. Polymer 2023, 285 , 126370. https://doi.org/10.1016/j.polymer.2023.126370
  54. Boyang Guo, Ximena Lopez‐Lorenzo, Yuan Fang, Eva Bäckström, Antonio Jose Capezza, Sudarsan Reddy Vanga, István Furó, Minna Hakkarainen, Per‐Olof Syrén. Fast Depolymerization of PET Bottle Mediated by Microwave Pre‐Treatment and An Engineered PETase**. ChemSusChem 2023, 16 (18) https://doi.org/10.1002/cssc.202300742
  55. Sean Najmi, Brandon C. Vance, Esun Selvam, Dylan Huang, Dionisios G. Vlachos. Controlling PET oligomers vs monomers via microwave-induced heating and swelling. Chemical Engineering Journal 2023, 471 , 144712. https://doi.org/10.1016/j.cej.2023.144712
  56. Ximena Lopez-Lorenzo, David Hueting, Eliott Bosshard, Per-Olof Syrén. WITHDRAWN: Degradation of PET microplastic particles to monomers in human serum by PETase. 2023https://doi.org/10.21203/rs.3.rs-3164368/v1
  57. Thore B. Thomsen, Sune Schubert, Cameron J. Hunt, Kim Borch, Kenneth Jensen, Jesper Brask, Peter Westh, Anne S. Meyer. Rate Response of Poly(Ethylene Terephthalate)‐Hydrolases to Substrate Crystallinity: Basis for Understanding the Lag Phase. ChemSusChem 2023, 16 (13) https://doi.org/10.1002/cssc.202300291
  58. José Augusto Castro-Rodríguez, Rogelio Rodríguez-Sotres, Amelia Farrés. Determinants for an Efficient Enzymatic Catalysis in Poly(Ethylene Terephthalate) Degradation. Catalysts 2023, 13 (3) , 591. https://doi.org/10.3390/catal13030591
  59. Junjie Huang, Dongxia Yan, Qingqing Zhu, Xiujie Cheng, Jing Tang, Xingmei Lu, Jiayu Xin. Depolymerization of polyethylene terephthalate with glycol under comparatively mild conditions. Polymer Degradation and Stability 2023, 208 , 110245. https://doi.org/10.1016/j.polymdegradstab.2022.110245
  60. Marco Orlando, Gianluca Molla, Pietro Castellani, Valentina Pirillo, Vincenzo Torretta, Navarro Ferronato. Microbial Enzyme Biotechnology to Reach Plastic Waste Circularity: Current Status, Problems and Perspectives. International Journal of Molecular Sciences 2023, 24 (4) , 3877. https://doi.org/10.3390/ijms24043877
  61. E. N. Efremenko, I. V. Lyagin, O. V. Maslova, O. V. Senko, N. A. Stepanov, A. G G. Aslanli. Catalytic degradation of microplastics. Russian Chemical Reviews 2023, 92 (2) , RCR5069. https://doi.org/10.57634/RCR5069
  62. Thore Bach Thomsen, Sune W. Schubert, Cameron J. Hunt, Peter Westh, Anne S. Meyer. A new continuous assay for quantitative assessment of enzymatic degradation of poly(ethylene terephthalate) (PET). Enzyme and Microbial Technology 2023, 162 , 110142. https://doi.org/10.1016/j.enzmictec.2022.110142
  63. Matthew Colachis, Nathan Clark, Ashley M. Frank, Edward B. Trigg, Colin Hinton, Greg Gregoriades, Ryan Daly, Vance Gustin, Bryon Moore, Rachel Thurston, Katarzyna H. Kucharzyk, Jacob L. Lilly. An Efficient and Scalable Melt Fiber Spinning System to Improve Enzyme-Based Pet Recycling. 2023https://doi.org/10.2139/ssrn.4672661
  64. Xing Zhou, Qi Wang, Sai Feng, Jingrui Deng, Keming Zhu, Yun Xing, Xiaolian Meng, Xiaojun Wang, Lu Li. Recycling Carbon Resources from Waste PET to Reduce Carbon Dioxide Emission: Carbonization Technology Review and Perspective. Journal of Renewable Materials 2023, 11 (5) , 2085-2108. https://doi.org/10.32604/jrm.2023.025032
  65. Natalia A. Tarazona, Ren Wei, Stefan Brott, Lara Pfaff, Uwe T. Bornscheuer, Andreas Lendlein, Rainhard Machatschek. Rapid depolymerization of poly(ethylene terephthalate) thin films by a dual-enzyme system and its impact on material properties. Chem Catalysis 2022, 2 (12) , 3573-3589. https://doi.org/10.1016/j.checat.2022.11.004
  66. Erika Erickson, Japheth E. Gado, Luisana Avilán, Felicia Bratti, Richard K. Brizendine, Paul A. Cox, Raj Gill, Rosie Graham, Dong-Jin Kim, Gerhard König, William E. Michener, Saroj Poudel, Kelsey J. Ramirez, Thomas J. Shakespeare, Michael Zahn, Eric S. Boyd, Christina M. Payne, Jennifer L. DuBois, Andrew R. Pickford, Gregg T. Beckham, John E. McGeehan. Sourcing thermotolerant poly(ethylene terephthalate) hydrolase scaffolds from natural diversity. Nature Communications 2022, 13 (1) https://doi.org/10.1038/s41467-022-35237-x
  67. Fusako Kawai, Yoshitomo Furushima, Norihiro Mochizuki, Naoki Muraki, Mitsuaki Yamashita, Akira Iida, Rie Mamoto, Takehiko Tosha, Ryo Iizuka, Sakihito Kitajima. Efficient depolymerization of polyethylene terephthalate (PET) and polyethylene furanoate by engineered PET hydrolase Cut190. AMB Express 2022, 12 (1) https://doi.org/10.1186/s13568-022-01474-y
  68. Josephine Herbert, Angela H. Beckett, Samuel C. Robson. A Review of Cross-Disciplinary Approaches for the Identification of Novel Industrially Relevant Plastic-Degrading Enzymes. Sustainability 2022, 14 (23) , 15898. https://doi.org/10.3390/su142315898
  69. Arpita Mrigwani, Bhishem Thakur, Purnananda Guptasarma. Conversion of polyethylene terephthalate into pure terephthalic acid through synergy between a solid-degrading cutinase and a reaction intermediate-hydrolysing carboxylesterase. Green Chemistry 2022, 24 (17) , 6707-6719. https://doi.org/10.1039/D2GC01965E

ACS Sustainable Chemistry & Engineering

Cite this: ACS Sustainable Chem. Eng. 2022, 10, 28, 9131–9140
Click to copy citationCitation copied!
https://doi.org/10.1021/acssuschemeng.2c01961
Published July 7, 2022

Copyright © 2022 The Authors. Published by American Chemical Society. This publication is licensed under

CC-BY 4.0 .

Article Views

6670

Altmetric

-

Citations

Learn about these metrics

Article Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.

Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.

The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated.

  • Abstract

    Figure 1

    Figure 1. Size distributions of CM-PET and HC-PET particles. Histograms indicate the volume percentage as a function of Feret’s minimum diameter of CM-PET (blue) and HC-PET (red) substrates. (A,D) 250, (B,E) 125, and (C,F) sub125 μm sieve fractions. Inset graphs show the expanded view of the data.

    Figure 2

    Figure 2. Monomer release as a function of time for 10 mg mL–1 PET particles with 1 μM LCC-ICCG. Monomers present in the reaction were analyzed by HPLC. (A) 250 μm CM-PET, (B) 125 μm CM-PET, (C) sub125 μm CM-PET, (D) 250 μm HC-PET, (E) 125 μm HC-PET, and (F) sub125 μm HC-PET. Reactions were performed in triplicate and errors bars represent the standard deviation. The data shown in this figure are provided in Dataset S1.

    Figure 3

    Figure 3. Total amount of monomers released from PET particles hydrolyzed with LCC-ICCG. The bars show the concentrations of all monomers released at the 72 h endpoint of the reaction with indicated [LCC-ICCG] and 10 mg mL–1 PET substrate. Reactions were performed in triplicate, and error bars represent standard deviation. The data shown in this figure are provided in Dataset S3.

    Figure 4

    Figure 4. InvMM kinetic analysis and total amount of monomers released from PET particles upon hydrolysis with LCC-ICCG. InvMM analysis of 10 mg mL–1 (A) CM-PET and (B) HC-PET hydrolysis rate as a function of LCC-ICCG concentration ([E]). Lines are fits of the invMM equation. Table 2 shows the fitted parameters. Reactions were performed in triplicate and error bars represent standard deviation. The data in this figure are provided in Dataset S5.

    Figure 5

    Figure 5. Conversion time courses of 1 L scale reactions conducted in bioreactors. Results from the reaction with various PET substrates at 100 g L–1 and 3 mg g–1 PET LCC-ICCG enzyme. (A–C) Product release profiles as a function of time measured by HPLC for (A) LC-PET film cut into ∼1 × 1 cm squares, (B) HC-PET particles, and (C) WC-PET particles. (D–F) Percent conversion of total PET for (D) LC-PET film cut into ∼1 × 1 cm squares, (E) HC-PET particles, and (F) WC-PET particles. Reactions were performed in duplicate, data points show the mean, and error bars show the absolute difference between the duplicates. The data shown in this figure are provided in Dataset S6.

  • References


    This article references 66 other publications.

    1. 1
      Cózar, A.; Echevarría, F.; González-Gordillo, J. I.; Irigoien, X.; Úbeda, B.; Hernández-León, S.; Palma, Á. T.; Navarro, S.; García-de-Lomas, J.; Ruiz, A. Plastic debris in the open ocean. Proc. Natl. Acad. Sci. U.S.A. 2014, 111, 1023910244,  DOI: 10.1073/pnas.1314705111
    2. 2
      Jambeck, J. R.; Geyer, R.; Wilcox, C.; Siegler, T. R.; Perryman, M.; Andrady, A.; Narayan, R.; Law, K. L. Marine pollution. Plastic waste inputs from land into the ocean. Science 2015, 347, 768771,  DOI: 10.1126/science.1260352
    3. 3
      Allen, S.; Allen, D.; Phoenix, V. R.; Le Roux, G.; Durántez Jiménez, P.; Simonneau, A.; Binet, S.; Galop, D. Atmospheric transport and deposition of microplastics in a remote mountain catchment. Nat. Geosci. 2019, 12, 339344,  DOI: 10.1038/s41561-019-0335-5
    4. 4
      Ellis, L. D.; Rorrer, N. A.; Sullivan, K. P.; Otto, M.; McGeehan, J. E.; Román-Leshkov, Y.; Wierckx, N.; Beckham, G. T. Chemical and biological catalysis for plastics recycling and upcycling. Nat. Catal. 2021, 4, 539556,  DOI: 10.1038/s41929-021-00648-4
    5. 5
      Nicholson, S. R.; Rorrer, N. A.; Carpenter, A. C.; Beckham, G. T. Manufacturing energy and greenhouse gas emissions associated with plastics consumption. Joule 2021, 5, 673686,  DOI: 10.1016/j.joule.2020.12.027
    6. 6
      PET Polymer: Chemical Economics Handbook; IHS Markit. https://ihsmarkit.com/products/pet-polymer-chemical-economics-handbook.html (accessed Sept 01, 2021).
    7. 7
      Ragaert, K.; Delva, L.; Van Geem, K. Mechanical and chemical recycling of solid plastic waste. Waste Manage. 2017, 69, 2458,  DOI: 10.1016/j.wasman.2017.07.044
    8. 8
      Sinha, V.; Patel, M. R.; Patel, J. V. Pet Waste Management by Chemical Recycling: A Review. J. Polym. Environ. 2010, 18, 825,  DOI: 10.1007/s10924-008-0106-7
    9. 9
      Wei, R.; Tiso, T.; Bertling, J.; O’Connor, K.; Blank, L. M.; Bornscheuer, U. T. Possibilities and limitations of biotechnological plastic degradation and recycling. Nat. Catal. 2020, 3, 867871,  DOI: 10.1038/s41929-020-00521-w
    10. 10
      Carniel, A.; Waldow, V. d. A.; Castro, A. M. d. A comprehensive and critical review on key elements to implement enzymatic PET depolymerization for recycling purposes. Biotechnol. Adv. 2021, 52, 107811,  DOI: 10.1016/j.biotechadv.2021.107811
    11. 11
      Wei, R.; Zimmermann, W. Biocatalysis as a green route for recycling the recalcitrant plastic polyethylene terephthalate. Microbiol. Biotechnol. 2017, 10, 13021307,  DOI: 10.1111/1751-7915.12714
    12. 12
      Müller, R.-J.; Schrader, H.; Profe, J.; Dresler, K.; Deckwer, W.-D. Enzymatic Degradation of Poly(ethylene terephthalate): Rapid Hydrolyse using a Hydrolase from T. fusca. Macromol. Rapid Commun. 2005, 26, 14001405,  DOI: 10.1002/marc.200500410
    13. 13
      Alisch-Mark, M.; Herrmann, A.; Zimmermann, W. Increase of the hydrophilicity of polyethylene terephthalate fibres by hydrolases from Thermomonospora fusca and Fusarium solani f. sp. pisi. Biotechnol. Lett. 2006, 28, 681685,  DOI: 10.1007/s10529-006-9041-7
    14. 14
      Eberl, A.; Heumann, S.; Brückner, T.; Araujo, R.; Cavaco-Paulo, A.; Kaufmann, F.; Kroutil, W.; Guebitz, G. M. Enzymatic surface hydrolysis of poly(ethylene terephthalate) and bis(benzoyloxyethyl) terephthalate by lipase and cutinase in the presence of surface active molecules. J. Biotechnol. 2009, 143, 207212,  DOI: 10.1016/j.jbiotec.2009.07.008
    15. 15
      Ronkvist, Å. M.; Xie, W.; Lu, W.; Gross, R. A. Cutinase-Catalyzed Hydrolysis of Poly(ethylene terephthalate). Macromolecules 2009, 42, 51285138,  DOI: 10.1021/ma9005318
    16. 16
      Ribitsch, D.; Heumann, S.; Trotscha, E.; Herrero Acero, E.; Greimel, K.; Leber, R.; Birner-Gruenberger, R.; Deller, S.; Eiteljoerg, I.; Remler, P. Hydrolysis of polyethyleneterephthalate by p-nitrobenzylesterase from Bacillus subtilis. Biotechnol. Prog. 2011, 27, 951960,  DOI: 10.1002/btpr.610
    17. 17
      Sulaiman, S.; Yamato, S.; Kanaya, E.; Kim, J.-J.; Koga, Y.; Takano, K.; Kanaya, S. Isolation of a novel cutinase homolog with polyethylene terephthalate-degrading activity from leaf-branch compost by using a metagenomic approach. Appl. Environ. Microbiol. 2012, 78, 15561562,  DOI: 10.1128/aem.06725-11
    18. 18
      Roth, C.; Wei, R.; Oeser, T.; Then, J.; Föllner, C.; Zimmermann, W.; Sträter, N. Structural and functional studies on a thermostable polyethylene terephthalate degrading hydrolase from Thermobifida fusca. Appl. Microbiol. Biotechnol. 2014, 98, 78157823,  DOI: 10.1007/s00253-014-5672-0
    19. 19
      Sulaiman, S.; You, D.-J.; Kanaya, E.; Koga, Y.; Kanaya, S. Crystal Structure and Thermodynamic and Kinetic Stability of Metagenome-Derived LC-Cutinase. Biochemistry 2014, 53, 18581869,  DOI: 10.1021/bi401561p
    20. 20
      Wei, R.; Oeser, T.; Zimmermann, W. Synthetic polyester-hydrolyzing enzymes from thermophilic actinomycetes. Adv. Appl. Microbiol. 2014, 89, 267305,  DOI: 10.1016/B978-0-12-800259-9.00007-X
    21. 21
      Perz, V.; Bleymaier, K.; Sinkel, C.; Kueper, U.; Bonnekessel, M.; Ribitsch, D.; Guebitz, G. M. Substrate specificities of cutinases on aliphatic-aromatic polyesters and on their model substrates. New Biotechnol. 2016, 33, 295304,  DOI: 10.1016/j.nbt.2015.11.004
    22. 22
      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, 11961199,  DOI: 10.1126/science.aad6359
    23. 23
      Ribitsch, D.; Hromic, A.; Zitzenbacher, S.; Zartl, B.; Gamerith, C.; Pellis, A.; Jungbauer, A.; Łyskowski, A.; Steinkellner, G.; Gruber, K. Small cause, large effect: Structural characterization of cutinases from Thermobifida cellulosilytica. Biotechnol. Bioeng. 2017, 114, 24812488,  DOI: 10.1002/bit.26372
    24. 24
      Kawai, F.; Kawabata, T.; Oda, M. Current State and Perspectives Related to the Polyethylene Terephthalate Hydrolases Available for Biorecycling. ACS Sustainable Chem. Eng. 2020, 8, 88948908,  DOI: 10.1021/acssuschemeng.0c01638
    25. 25
      Sonnendecker, C.; Oeser, J.; Richter, P. K.; Hille, P.; Zhao, Z.; Fischer, C.; Lippold, H.; Blazquez-Sanchez, P.; Engelberger, F.; Ramirez-Sarmiento, C. A. Low Carbon Footprint Recycling of Post-Consumer PET Plastic with a Metagenomic Polyester Hydrolase. ChemSusChem 2022, 15, e202101062  DOI: 10.1002/cssc.202101062
    26. 26
      Herrero Acero, E.; Ribitsch, D.; Dellacher, A.; Zitzenbacher, S.; Marold, A.; Steinkellner, G.; Gruber, K.; Schwab, H.; Guebitz, G. M. Surface engineering of a cutinase from Thermobifida cellulosilytica for improved polyester hydrolysis. Biotechnol. Bioeng. 2013, 110, 25812590,  DOI: 10.1002/bit.24930
    27. 27
      Ribitsch, D.; Yebra, A. O.; Zitzenbacher, S.; Wu, J.; Nowitsch, S.; Steinkellner, G.; Greimel, K.; Doliska, A.; Oberdorfer, G.; Gruber, C. C. Fusion of binding domains to Thermobifida cellulosilytica cutinase to tune sorption characteristics and enhancing PET hydrolysis. Biomacromolecules 2013, 14, 17691776,  DOI: 10.1021/bm400140u
    28. 28
      Ribitsch, D.; Herrero Acero, E.; Przylucka, A.; Zitzenbacher, S.; Marold, A.; Gamerith, C.; Tscheliessnig, R.; Jungbauer, A.; Rennhofer, H.; Lichtenegger, H. Enhanced cutinase-catalyzed hydrolysis of polyethylene terephthalate by covalent fusion to hydrophobins. Appl. Environ. Microbiol. 2015, 81, 35863592,  DOI: 10.1128/AEM.04111-14
    29. 29
      Then, J.; Wei, R.; Oeser, T.; Barth, M.; Belisário-Ferrari, M. R.; Schmidt, J.; Zimmermann, W. Ca2+ and Mg2+ binding site engineering increases the degradation of polyethylene terephthalate films by polyester hydrolases from Thermobifida fusca. Biotechnol. J. 2015, 10, 592598,  DOI: 10.1002/biot.201400620
    30. 30
      Wei, R.; Oeser, T.; Schmidt, J.; Meier, R.; Barth, M.; Then, J.; Zimmermann, W. Engineered bacterial polyester hydrolases efficiently degrade polyethylene terephthalate due to relieved product inhibition. Biotechnol. Bioeng. 2016, 113, 16581665,  DOI: 10.1002/bit.25941
    31. 31
      Shirke, A. N.; Basore, D.; Butterfoss, G. L.; Bonneau, R.; Bystroff, C.; Gross, R. A. Toward rational thermostabilization of Aspergillus oryzae cutinase: Insights into catalytic and structural stability. Proteins 2016, 84, 6072,  DOI: 10.1002/prot.24955
    32. 32
      Biundo, A.; Ribitsch, D.; Steinkellner, G.; Gruber, K.; Guebitz, G. M. Polyester hydrolysis is enhanced by a truncated esterase: Less is more. Biotechnol. J. 2017, 12,  DOI: 10.1002/biot.201600450
    33. 33
      Shirke, A. N.; Butterfoss, G. L.; Saikia, R.; Basu, A.; Maria, L.; Svendsen, A.; Gross, R. A. Engineered Humicola insolens cutinase for efficient cellulose acetate deacetylation. Biotechnol. J. 2017, 12, 1700188,  DOI: 10.1002/biot.201700188
    34. 34
      Austin, H. P.; Allen, M. D.; Donohoe, B. S.; Rorrer, N. A.; Kearns, F. L.; Silveira, R. L.; Pollard, B. C.; Dominick, G.; Duman, R.; El Omari, K. Characterization and engineering of a plastic-degrading aromatic polyesterase. Proc. Natl. Acad. Sci. U.S.A. 2018, 115, E4350E4357,  DOI: 10.1073/pnas.1718804115
    35. 35
      Biundo, A.; Ribitsch, D.; Guebitz, G. M. Surface engineering of polyester-degrading enzymes to improve efficiency and tune specificity. Appl. Microbiol. Biotechnol. 2018, 102, 35513559,  DOI: 10.1007/s00253-018-8850-7
    36. 36
      Shirke, A. N.; White, C.; Englaender, J. A.; Zwarycz, A.; Butterfoss, G. L.; Linhardt, R. J.; Gross, R. A. Stabilizing Leaf and Branch Compost Cutinase (LCC) with Glycosylation: Mechanism and Effect on PET Hydrolysis. Biochemistry 2018, 57, 11901200,  DOI: 10.1021/acs.biochem.7b01189
    37. 37
      Son, H. F.; Cho, I. J.; Joo, S.; Seo, H.; Sagong, H.-Y.; Choi, S. Y.; Lee, S. Y.; Kim, K.-J. Rational Protein Engineering of Thermo-Stable PETase from Ideonella sakaiensis for Highly Efficient PET Degradation. ACS Catal. 2019, 9, 35193526,  DOI: 10.1021/acscatal.9b00568
    38. 38
      Furukawa, M.; Kawakami, N.; Tomizawa, A.; Miyamoto, K. Efficient Degradation of Poly(ethylene terephthalate) with Thermobifida fusca Cutinase Exhibiting Improved Catalytic Activity Generated using Mutagenesis and Additive-based Approaches. Sci. Rep. 2019, 9, 16038,  DOI: 10.1038/s41598-019-52379-z
    39. 39
      Tournier, V.; Topham, C. M.; Gilles, A.; David, B.; Folgoas, C.; Moya-Leclair, E.; Kamionka, E.; Desrousseaux, M.-L.; Texier, H.; Gavalda, S. An engineered PET depolymerase to break down and recycle plastic bottles. Nature 2020, 580, 216219,  DOI: 10.1038/s41586-020-2149-4
    40. 40
      Bell, E.; Smithson, R.; Kilbride, S.; Foster, J.; Hardy, F.; Ramachandran, S.; Tedstone, A.; Haigh, S.; Garforth, A.; Day, P. Directed Evolution of an Efficient and Thermostable PET Depolymerase. ChemRxiv 2021,  DOI: 10.26434/chemrxiv-2021-mcjh6
    41. 41
      Cui, Y.; Chen, Y.; Liu, X.; Dong, S.; Tian, Y. e.; Qiao, Y.; Mitra, R.; Han, J.; Li, C.; Han, X. Computational Redesign of a PETase for Plastic Biodegradation under Ambient Condition by the GRAPE Strategy. ACS Catal. 2021, 11, 13401350,  DOI: 10.1021/acscatal.0c05126
    42. 42
      Nakamura, A.; Kobayashi, N.; Koga, N.; Iino, R. Positive Charge Introduction on the Surface of Thermostabilized PET Hydrolase Facilitates PET Binding and Degradation. ACS Catal. 2021, 11, 85508564,  DOI: 10.1021/acscatal.1c01204
    43. 43
      Guo, B.; Vanga, S. R.; Lopez-Lorenzo, X.; Saenz-Mendez, P.; Ericsson, S. R.; Fang, Y.; Ye, X.; Schriever, K.; Bäckström, E.; Biundo, A. Conformational Selection in Biocatalytic Plastic Degradation by PETase. ACS Catal. 2022, 12, 33973409,  DOI: 10.1021/acscatal.1c05548
    44. 44
      Lu, H.; Diaz, D. J.; Czarnecki, N. J.; Zhu, C.; Kim, W.; Shroff, R.; Acosta, D. J.; Alexander, B. R.; Cole, H. O.; Zhang, Y. Machine learning-aided engineering of hydrolases for PET depolymerization. Nature 2022, 604, 662667,  DOI: 10.1038/s41586-022-04599-z
    45. 45
      Wei, R.; von Haugwitz, G.; Pfaff, L.; Mican, J.; Badenhorst, C. P. S.; Liu, W.; Weber, G.; Austin, H. P.; Bednar, D.; Damborsky, J. Mechanism-Based Design of Efficient PET Hydrolases. ACS Catal. 2022, 12, 33823396,  DOI: 10.1021/acscatal.1c05856
    46. 46
      Zeng, W.; Li, X.; Yang, Y.; Min, J.; Huang, J.-W.; Liu, W.; Niu, D.; Yang, X.; Han, X.; Zhang, L. Substrate-Binding Mode of a Thermophilic PET Hydrolase and Engineering the Enzyme to Enhance the Hydrolytic Efficacy. ACS Catal. 2022, 12, 30333040,  DOI: 10.1021/acscatal.1c05800
    47. 47
      Singh, A.; Rorrer, N. A.; Nicholson, S. R.; Erickson, E.; DesVeaux, J. S.; Avelino, A. F. T.; Lamers, P.; Bhatt, A.; Zhang, Y.; Avery, G. Techno-economic, life-cycle, and socioeconomic impact analysis of enzymatic recycling of poly(ethylene terephthalate). Joule 2021, 5, 24792503,  DOI: 10.1016/j.joule.2021.06.015
    48. 48
      Wei, R.; Breite, D.; Song, C.; Gräsing, D.; Ploss, T.; Hille, P.; Schwerdtfeger, R.; Matysik, J.; Schulze, A.; Zimmermann, W. Biocatalytic Degradation Efficiency of Postconsumer Polyethylene Terephthalate Packaging Determined by Their Polymer Microstructures. Adv. Sci. 2019, 6, 1900491,  DOI: 10.1002/advs.201900491
    49. 49
      Knott, B. C.; Erickson, E.; Allen, M. D.; Gado, J. E.; Graham, R.; Kearns, F. L.; Pardo, I.; Topuzlu, E.; Anderson, J. J.; Austin, H. P. Characterization and engineering of a two-enzyme system for plastics depolymerization. Proc. Natl. Acad. Sci. U.S.A. 2020, 117, 2547625485,  DOI: 10.1073/pnas.2006753117
    50. 50
      Erickson, E.; Shakespeare, T. J.; Bratti, F.; Buss, B. L.; Graham, R.; Hawkins, M. A.; König, G.; Michener, W. E.; Miscall, J.; Ramirez, K. J. Comparative performance of PETase as a function of reaction conditions, substrate properties, and product accumulation. ChemSusChem 2022, 15, e202101932  DOI: 10.1002/cssc.202101932
    51. 51
      Lu, H.; Diaz, D. J.; Czarnecki, N. J.; Zhu, C.; Kim, W.; Shroff, R.; Acosta, D. J.; Alexander, B.; Cole, H.; Zhang, Y. J. Deep learning redesign of PETase for practical PET degrading applications. bioRxiv 2021,  DOI: 10.1101/2021.10.10.463845
    52. 52
      de Castro, A. M.; Carniel, A.; Nicomedes Junior, J.; da Conceição Gomes, A.; Valoni, É. Screening of commercial enzymes for poly(ethylene terephthalate) (PET) hydrolysis and synergy studies on different substrate sources. J. Ind. Microbiol. Biotechnol. 2017, 44, 835844,  DOI: 10.1007/s10295-017-1942-z
    53. 53
      Gamerith, C.; Zartl, B.; Pellis, A.; Guillamot, F.; Marty, A.; Acero, E. H.; Guebitz, G. M. Enzymatic recovery of polyester building blocks from polymer blends. Process Biochem. 2017, 59, 5864,  DOI: 10.1016/j.procbio.2017.01.004
    54. 54
      Castro, A. M. d.; Carniel, A.; Stahelin, D.; Chinelatto Junior, L. S.; Honorato, H. d. A.; de Menezes, S. M. C. High-fold improvement of assorted post-consumer poly(ethylene terephthalate) (PET) packages hydrolysis using Humicola insolens cutinase as a single biocatalyst. Process Biochem. 2019, 81, 8591,  DOI: 10.1016/j.procbio.2019.03.006
    55. 55
      Kari, J.; Andersen, M.; Borch, K.; Westh, P. An Inverse Michaelis–Menten Approach for Interfacial Enzyme Kinetics. ACS Catal. 2017, 7, 49044914,  DOI: 10.1021/acscatal.7b00838
    56. 56
      Andersen, M.; Kari, J.; Borch, K.; Westh, P. Michaelis-Menten equation for degradation of insoluble substrate. Math. Biosci. 2018, 296, 9397,  DOI: 10.1016/j.mbs.2017.11.011
    57. 57
      Bååth, J. A.; Borch, K.; Jensen, K.; Brask, J.; Westh, P. Comparative Biochemistry of Four Polyester (PET) Hydrolases. ChemBioChem 2021, 22, 16271637,  DOI: 10.1002/cbic.202000793
    58. 58
      Werner, A. Z.; Clare, R.; Mand, T. D.; Pardo, I.; Ramirez, K. J.; Haugen, S. J.; Bratti, F.; Dexter, G. N.; Elmore, J. R.; Huenemann, J. D. Tandem chemical deconstruction and biological upcycling of poly(ethylene terephthalate) to beta-ketoadipic acid by Pseudomonas putida KT2440. Metab. Eng. 2021, 67, 250261,  DOI: 10.1016/j.ymben.2021.07.005
    59. 59
      Blundell, D. J.; MacKerron, D. H.; Fuller, W.; Mahendrasingam, A.; Martin, C.; Oldman, R. J.; Rule, R. J.; Riekel, C. Characterization of strain-induced crystallization of poly(ethylene terephthalate) at fast draw rates using synchrotron radiation. Polymer 1996, 37, 33033311,  DOI: 10.1016/0032-3861(96)88476-X
    60. 60
      Blundell, D. J.; Mahendrasingam, A.; Martin, C.; Fuller, W.; MacKerron, D. H.; Harvie, J. L.; Oldman, R. J.; Riekel, C. Orientation prior to crystallisation during drawing of poly(ethylene terephthalate). Polymer 2000, 41, 77937802,  DOI: 10.1016/S0032-3861(00)00128-2
    61. 61
      Mahendrasingam, A.; Martin, C.; Fuller, W.; Blundell, D. J.; Oldman, R. J.; MacKerron, D. H.; Harvie, J. L.; Riekel, C. Observation of a transient structure prior to strain-induced crystallization in poly(ethylene terephthalate). Polymer 2000, 41, 12171221,  DOI: 10.1016/S0032-3861(99)00461-9
    62. 62
      Forestier, E.; Combeaud, C.; Guigo, N.; Sbirrazzuoli, N.; Billon, N. Understanding of strain-induced crystallization developments scenarios for polyesters: Comparison of poly(ethylene furanoate), PEF, and poly(ethylene terephthalate), PET. Polymer 2020, 203, 122755,  DOI: 10.1016/j.polymer.2020.122755
    63. 63
      Bashir, Z.; Al-Aloush, I.; Al-Raqibah, I.; Ibrahim, M. Evaluation of three methods for the measurement of crystallinity of PET resins, preforms, and bottles. Polym. Eng. Sci. 2000, 40, 24422455,  DOI: 10.1002/pen.11376
    64. 64
      Scandola, M.; Focarete, M. L.; Frisoni, G. Simple Kinetic Model for the Heterogeneous Enzymatic Hydrolysis of Natural Poly(3-hydroxybutyrate). Macromolecules 1998, 31, 38463851,  DOI: 10.1021/ma980137y
    65. 65
      Thomsen, T. B.; Hunt, C. J.; Meyer, A. S. Influence of substrate crystallinity and glass transition temperature on enzymatic degradation of polyethylene terephthalate (PET). New Biotechnol. 2022, 69, 2835,  DOI: 10.1016/j.nbt.2022.02.006
    66. 66
      Barth, M.; Honak, A.; Oeser, T.; Wei, R.; Belisário-Ferrari, M. R.; Then, J.; Schmidt, J.; Zimmermann, W. A dual enzyme system composed of a polyester hydrolase and a carboxylesterase enhances the biocatalytic degradation of polyethylene terephthalate films. Biotechnol. J. 2016, 11, 10821087,  DOI: 10.1002/biot.201600008
  • Supporting Information

    Supporting Information


    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acssuschemeng.2c01961.

    • Additional characterization and kinetic data as well as datasets for all figures (PDF)


    Terms & Conditions

    Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.