General features to enhance enzymatic activity of poly(ethylene terephthalate) hydrolysis

Chen, Chun‐Chi; Han, Xu; Li, Xian; Jiang, Pengcheng; Niu, Du; Ma, Lixin; Liu, Weidong; Li, Siyu; Qu, Yingying; Hu, Hongliang; Min, Jian; Yang, Yu; Zhang, Lilan; Zeng, Wei; Huang, Jian‐Wen; Dai, Longhai; Guo, Rey‐Ting

doi:10.1038/s41929-021-00616-y

Cited by 136 publications

(173 citation statements)

References 30 publications

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“…8 Here, especially the known structures of the PETase from Ideonella sakaiensis (for example PDB code 5XH3; Han et al, 2017 ), show high consistency indicated by the rmsd of 1.3-1.5 Å. Also, high structural similarity can be seen with the structure of PE-H from Pseudomonas aestusnigri (PDB code 6SCD rmsd 1.3) and the cutinase from a member of the Burkhoderiales bacteria family (PDB code 7CWQ, rmsd of 1.32 ( Bollinger et al, 2020 ; Chen et al, 2021 ). One disulfide bond is present in PET30ΔPorC (C262-C285), a common feature for Type II PET-degrading enzymes ( Joo et al, 2018 ).…”

Section: Resultsmentioning

confidence: 85%

The Bacteroidetes Aequorivita sp. and Kaistella jeonii Produce Promiscuous Esterases With PET-Hydrolyzing Activity

et al. 2022

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Certain members of the Actinobacteria and Proteobacteria are known to degrade polyethylene terephthalate (PET). Here, we describe the first functional PET-active enzymes from the Bacteroidetes phylum. Using a PETase-specific Hidden-Markov-Model- (HMM-) based search algorithm, we identified several PETase candidates from Flavobacteriaceae and Porphyromonadaceae. Among them, two promiscuous and cold-active esterases derived from Aequorivita sp. (PET27) and Kaistella jeonii (PET30) showed depolymerizing activity on polycaprolactone (PCL), amorphous PET foil and on the polyester polyurethane Impranil® DLN. PET27 is a 37.8 kDa enzyme that released an average of 174.4 nmol terephthalic acid (TPA) after 120 h at 30°C from a 7 mg PET foil platelet in a 200 μl reaction volume, 38-times more than PET30 (37.4 kDa) released under the same conditions. The crystal structure of PET30 without its C-terminal Por-domain (PET30ΔPorC) was solved at 2.1 Å and displays high structural similarity to the IsPETase. PET30 shows a Phe-Met-Tyr substrate binding motif, which seems to be a unique feature, as IsPETase, LCC and PET2 all contain Tyr-Met-Trp binding residues, while PET27 possesses a Phe-Met-Trp motif that is identical to Cut190. Microscopic analyses showed that K. jeonii cells are indeed able to bind on and colonize PET surfaces after a few days of incubation. Homologs of PET27 and PET30 were detected in metagenomes, predominantly aquatic habitats, encompassing a wide range of different global climate zones and suggesting a hitherto unknown influence of this bacterial phylum on man-made polymer degradation.

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Section: Resultsmentioning

confidence: 85%

The Bacteroidetes Aequorivita sp. and Kaistella jeonii Produce Promiscuous Esterases With PET-Hydrolyzing Activity

et al. 2022

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“…For example, an engineered leaf-branch compost cutinase (LCC) can degrade 90% of pretreated pc-PET within 10 hours at 72 ºC and a pH of 8.0 4 . Most other PHEs similarly show poor activity at moderate temperatures 9 and more neutral pH conditions 10 , greatly restricting in situ / microbiallyenabled degradation solutions for PET waste. This limitation is of critical concern as 40% of uncollectable plastics reside in natural environments 11 .…”

Section: Manuscript Textmentioning

confidence: 99%

Deep learning redesign of PETase for practical PET degrading applications

Diaz

Czarnecki

et al. 2021

Preprint

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Plastic waste poses an ecological challenge. While current plastic waste management largely relies on unsustainable, energy-intensive, or even hazardous physicochemical and mechanical processes, enzymatic degradation offers a green and sustainable route for plastic waste recycling. Poly(ethylene terephthalate) (PET) has been extensively used in packaging and for the manufacture of fabrics and single-used containers, accounting for 12% of global solid waste. The practical application of PET hydrolases has been hampered by their lack of robustness and the requirement for high processing temperatures. Here, we use a structure-based, deep learning algorithm to engineer an extremely robust and highly active PET hydrolase. Our best resulting mutant (FAST-PETase: Functional, Active, Stable, and Tolerant PETase) exhibits superior PET-hydrolytic activity relative to both wild-type and engineered alternatives, (including a leaf-branch compost cutinase and its mutant) and possesses enhanced thermostability and pH tolerance. We demonstrate that whole, untreated, post-consumer PET from 51 different plastic products can all be completely degraded by FAST-PETase within one week, and in as little as 24 hours at 50 C. Finally, we demonstrate two paths for closed-loop PET recycling and valorization. First, we re-synthesize virgin PET from the monomers recovered after enzymatic depolymerization. Second, we enable in situ microbially-enabled valorization using a Pseudomonas strain together with FAST-PETase to degrade PET and utilize the evolved monomers as a carbon source for growth and polyhydroxyalkanoate production. Collectively, our results demonstrate the substantial improvements enabled by deep learning and a viable route for enzymatic plastic recycling at the industrial scale.

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“…PET-hydrolytic activity was measured as previously described ( Espino-Rammer et al, 2013 ; Chen et al, 2021 ; Kaabel et al, 2021 ). Briefly, 10 μg/ml purified enzyme was incubated with 4 mg high-crystallinity PET microparticles (42.67% crystallinity) in 20 mM Tris-HCl (pH 8.5), 500 mM NaCl, 10% ( v / v ) dimethyl sulfoxide, and 10 mM CaCl 2 .…”

Section: Methodsmentioning

confidence: 99%

Catalytic Features and Thermal Adaptation Mechanisms of a Deep Sea Bacterial Cutinase-Type Poly(Ethylene Terephthalate) Hydrolase

Liu

et al. 2022

Front. Bioeng. Biotechnol.

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Poly (ethylene terephthalate) (PET) plastic is chemically inert and persistent. Massive quantities of PET waste end up in landfill sites and oceans, posing major global pollution concerns. PET degrading enzymes with high efficiency provide plastic recycling and bioremediation possibilities. Here, we report a novel cutinase, MtCut with distinct catalytic behaviors, derived from the deep sea Nocardiopsaceae family strain. Biochemical analyses showed MtCut efficiently hydrolyzed PET at ambient temperatures and in an exo-type manner. The activity and stability of MtCut were enhanced by the addition of calcium ions. Notably, no hydrolysis products inhibition was observed during PET depolymerization, suggesting MtCut is a better biocatalyst when compared to other PET hydrolases. In addition, structural components associated with thermal adaptation were investigated using molecular dynamic (MD) simulations, and key regions regulating MtCut thermostability were identified. Our biochemical and structural analyses of MtCut deepen the understanding of PET hydrolysis by cutinases, and provide invaluable insights on improvement and performance engineering strategies for PET-degrading biocatalysts.

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General features to enhance enzymatic activity of poly(ethylene terephthalate) hydrolysis

Cited by 136 publications

References 30 publications

The Bacteroidetes Aequorivita sp. and Kaistella jeonii Produce Promiscuous Esterases With PET-Hydrolyzing Activity

The Bacteroidetes Aequorivita sp. and Kaistella jeonii Produce Promiscuous Esterases With PET-Hydrolyzing Activity

Deep learning redesign of PETase for practical PET degrading applications

Catalytic Features and Thermal Adaptation Mechanisms of a Deep Sea Bacterial Cutinase-Type Poly(Ethylene Terephthalate) Hydrolase

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