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      Efficient Depolymerization of Poly(ethylene 2,5-furanoate) Using Polyester Hydrolases

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          Abstract

          Poly(ethylene 2,5-furanoate) (PEF) is considered to be the next-generation green polyester and is hailed as a rising star among novel plastics. It is biobased, is nontoxic, and has comparable or improved properties compared to polyethylene terephthalate (PET). Biobased PEF offers lower life-cycle greenhouse gas emissions than PET. However, with its industrial production starting soon, relatively little is known about its actual recyclability. This work reports on the near complete depolymerization of PEF using two efficient PET hydrolases, FastPETase and leaf compost-cutinase (LCC), at loadings 4.5–17 times lower than previously reported. FastPETase and LCC exhibited maximum depolymerization of PEF, measured by weight loss and 2,5-furandicarboxylic acid (FDCA) production, using potassium phosphate–NaOH buffer at 50 and 65 °C, respectively. The 98% depolymerization of 13 g L –1 PEF film was achieved by three additions of the LCC in 72 h, while 78% weight loss was obtained using FastPETase in controlled conditions. Nonetheless, 92% weight loss was obtained with FastPETase when using only 6 g L –1 PEF. The main reaction products were identified as FDCA, ethylene glycol, and mono(2-hydroxyethyl)-furanoate. LCC performed better than FastPETase, in terms of both FDCA release and weight loss. The effect of crystallinity was evident on the enzymes’ performance, as only 4% to 7% weight loss of crystalline PEF (32%) was recorded. Microscopy studies of the treated PEF films provided information on the surface erosion processes and revealed higher resistance of the crystalline phase, explaining the low level of depolymerization. The study presents important insights into the enzymatic hydrolysis of biobased PEF material and paves the path toward more viable applications within biopolymer waste recycling.

          Abstract

          This study provides an insight into the enzyme-based hydrolysis of PEF, improving the potential for industrial enzymatic recycling of this promising bioplastic that soon will be available on the market.

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          Most cited references26

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          An engineered PET depolymerase to break down and recycle plastic bottles

          Present estimates suggest that of the 359 million tons of plastics produced annually worldwide1, 150-200 million tons accumulate in landfill or in the natural environment2. Poly(ethylene terephthalate) (PET) is the most abundant polyester plastic, with almost 70 million tons manufactured annually worldwide for use in textiles and packaging3. The main recycling process for PET, via thermomechanical means, results in a loss of mechanical properties4. Consequently, de novo synthesis is preferred and PET waste continues to accumulate. With a high ratio of aromatic terephthalate units-which reduce chain mobility-PET is a polyester that is extremely difficult to hydrolyse5. Several PET hydrolase enzymes have been reported, but show limited productivity6,7. Here we describe an improved PET hydrolase that ultimately achieves, over 10 hours, a minimum of 90 per cent PET depolymerization into monomers, with a productivity of 16.7 grams of terephthalate per litre per hour (200 grams per kilogram of PET suspension, with an enzyme concentration of 3 milligrams per gram of PET). This highly efficient, optimized enzyme outperforms all PET hydrolases reported so far, including an enzyme8,9 from the bacterium Ideonella sakaiensis strain 201-F6 (even assisted by a secondary enzyme10) and related improved variants11-14 that have attracted recent interest. We also show that biologically recycled PET exhibiting the same properties as petrochemical PET can be produced from enzymatically depolymerized PET waste, before being processed into bottles, thereby contributing towards the concept of a circular PET economy.
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            Isolation of a novel cutinase homolog with polyethylene terephthalate-degrading activity from leaf-branch compost by using a metagenomic approach.

            The gene encoding a cutinase homolog, LC-cutinase, was cloned from a fosmid library of a leaf-branch compost metagenome by functional screening using tributyrin agar plates. LC-cutinase shows the highest amino acid sequence identity of 59.7% to Thermomonospora curvata lipase. It also shows the 57.4% identity to Thermobifida fusca cutinase. When LC-cutinase without a putative signal peptide was secreted to the periplasm of Escherichia coli cells with the assistance of the pelB leader sequence, more than 50% of the recombinant protein, termed LC-cutinase*, was excreted into the extracellular medium. It was purified and characterized. LC-cutinase* hydrolyzed various fatty acid monoesters with acyl chain lengths of 2 to 18, with a preference for short-chain substrates (C(4) substrate at most) most optimally at pH 8.5 and 50°C, but could not hydrolyze olive oil. It lost activity with half-lives of 40 min at 70°C and 7 min at 80°C. LC-cutinase* had an ability to degrade poly(ε-caprolactone) and polyethylene terephthalate (PET). The specific PET-degrading activity of LC-cutinase* was determined to be 12 mg/h/mg of enzyme (2.7 mg/h/μkat of pNP-butyrate-degrading activity) at pH 8.0 and 50°C. This activity is higher than those of the bacterial and fungal cutinases reported thus far, suggesting that LC-cutinase* not only serves as a good model for understanding the molecular mechanism of PET-degrading enzyme but also is potentially applicable for surface modification and degradation of PET.
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              Machine learning-aided engineering of hydrolases for PET depolymerization

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                Author and article information

                Journal
                ACS Sustain Chem Eng
                ACS Sustain Chem Eng
                sc
                ascecg
                ACS Sustainable Chemistry & Engineering
                American Chemical Society
                2168-0485
                20 June 2024
                01 July 2024
                : 12
                : 26
                : 9658-9668
                Affiliations
                []Department of Chemistry and Bioscience, Aalborg University , Fredrik Bajers Vej 7H, 9220 Aalborg, Denmark
                []Dipartimento di Chimica e Chimica Industriale, Universitá degli Studi di Genova , Via Dodecaneso 31, Genova 16146, Italy
                [§ ]Department of Materials and Production, Aalborg University , Fibigerstræde 16, 9220 Aalborg, Denmark
                Author notes
                Author information
                https://orcid.org/0000-0001-6942-5056
                https://orcid.org/0000-0002-9480-3765
                https://orcid.org/0000-0002-5638-6390
                Article
                10.1021/acssuschemeng.4c00915
                11220789
                38966237
                3710aff7-73ac-494c-828b-3ae300030815
                © 2024 The Authors. Published by American Chemical Society

                Permits non-commercial access and re-use, provided that author attribution and integrity are maintained; but does not permit creation of adaptations or other derivative works ( https://creativecommons.org/licenses/by-nc-nd/4.0/).

                History
                : 31 January 2024
                : 03 June 2024
                : 01 June 2024
                Funding
                Funded by: H2020 LEIT Biotechnology, doi 10.13039/100010689;
                Award ID: 953073
                Categories
                Research Article
                Custom metadata
                sc4c00915
                sc4c00915

                pef,enzymatic depolymerization,bioplastics,plastic recycling,circular economy,polyester

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