JOURNAL OF HAZARDOUS MATERIALS
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2024
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IF 12.2
Si-Qi Chen,Qing-Song Huang,Yang Li,Jing Wu,Sheng Chen,Zheng-Fei Yan
Polyethylene terephthalate (PET) is one of the most widely used plastics, but its fragmentation into microplastics poses significant environmental challenges. The recycling of PET microplastics is hindered by their low solubility and widespread dispersion in the environment, making microbial in-situ degradation a promising solution. However, existing PET-degrading strains exhibited the limited effectiveness, primarily due to the diffusion of secreted hydrolases away from the PET surface. In this study, Stenotrophomonas pavanii JWG-G1 was engineered to achieve the targeted aggregation of PET hydrolase PETase on the cell surface by fusing it with an endogenous anchor protein. This approach aims to maximise the local concentration of PETase around PET, thereby increasing the overall rate of PET degradation. The PETase surface-aggregated system, S. pavanii /PaL-PETase, demonstrated the highest degradation efficiency, achieving 63.3 % degradation of low-crystallinity PET (lcPET) and 27.3 % degradation of high-crystallinity PET bottles (hcPET) at 30 °C. This represents the highest degradation rate reported for a displayed whole-cell system at ambient temperature. Furthermore, this system exhibited broad-spectrum degradation activity against various polyesters. These findings suggest that this system offers a promising, eco-friendly solution to PET and other polyester pollution, with potential implications for environmental bioremediation strategies.
研究领域:
环境科学
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微生物学
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生物技术
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高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2021
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IF 3.978
Kun Chen,Mingqin Quan,Xiaoyan Dong,Qinghong Shi,Yan Sun
Polyethylene terephthalate is one of the most widely used synthetic plastics. The accumulation of abandoned PET has brought an environmental problem. A recently reported PET hydrolase , PETase, displays an efficient decomposition to PET. Herein, we synthesized four PETase conjugates by modifying four types of monomers onto PETase, which were hydrophobic tert-butyl methacrylate (TBMA), hydrophilic hydroxyethyl methacrylate (HEMA), positively charged 2-(dimethylamino)ethyl methacrylate (DMAEMA), and negatively charged methacrylic acid (MA). Despite the low modification with an average number of 3–5 monomers on each enzyme molecule, all the PETase conjugates exhibited improved enzyme activity , and thermal and pH stability. Particularly, the catalytic efficiency ( k cat / K m ) of TBMA-/DMAEMA-PETase toward bis-2(hydroxyethyl) terephthalate (BHET) increased 1.4 and 1.6-fold as compared with PETase. Characterizations by fluorescence and circular dichroism spectroscopy unraveled that the modification induced the structural transformations, including compact overall structure and opened substrate-binding pocket, which were in favor of the enhanced catalytic performance. Biodegradation against PET films by TBMA-/DMAEMA-PETase were 4.69 and 3.26-fold higher than PETase. The different catalytic performances of the enzyme conjugates for BHET and PET are considered due to the different interactions of the conjugates with the different substrates. The work demonstrated that the low modifications with hydrophobic and hydrophilic monomers were both effective in improving the catalytic performance toward PET degradation, especially for the hydrophobic and positively charged monomers.
研究领域:
生物技术
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环境科学
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酶学
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材料科学
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高影响因子
ACS Applied Materials & Interfaces
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2024
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IF 8.3
Zhi Qu,Yan Sun
The enormous accumulation of poly(ethylene terephthalate) (PET) waste has posed a serious threat to the environment and human health, and biodegradation with PET hydrolase (PETase) can be a possible solution. Herein, we propose site-specifically modifying PETase with amphiphilic polymers to improve the enzyme performance at ambient temperature. For this purpose, we devise a computer-aided strategy to prioritize the conjugation site, and polyethylene glycol (PEG) preparations of 0.55 to 10 kDa are site-specifically conjugated to PETase. The most active conjugate PETase–PEG 5k (PETase-5K) shows an increase of melting temperature (3.88 °C) and significantly improves PET degradation performance (3.5- and 3.1-fold increases at 30 and 40 °C, respectively). Experimental investigation and molecular dynamics simulations reveal that the site-specific PEGylation increases the hydrophobic solvent-accessible surface area and the binding capability to the PET surface, thickens the hydration layer, increases the intramolecular hydrogen bonding, reduces the interactions between water and the conjugated enzyme surface, and rigidifies the enzyme structure via hydrogen bonding and hydrophobic interactions between the polymer and the enzyme, thus leading to improved enzymatic performance of PETase-5K. We further validate the versatility of the site-specific PEGylation in one of the most evolved variants of PETase, FAST-PETase, by 1.8-fold improvement in PET degradation at 30 °C. The presented computer-aided site-specific conjugation strategy has opened a new avenue to enhancing PETase performance at ambient temperature, and the contribution of PEGylation to PETase unraveled in this work laid a foundation for the rational engineering of PET hydrolases.
研究领域:
环境科学
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生物化学
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生物技术
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分子生物学
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材料科学
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高影响因子
JOURNAL OF HAZARDOUS MATERIALS
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2026
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IF 10.6
Zhi Qu,Zongyang Tian,Zehui Guo,Yan Sun
Extensive accumulation of polyethylene terephthalate (PET) plastic waste causes serious pollution to the global environment, and developing superior PET hydrolases is vital for the enzymatic degradation and biorecycling of PET. Here, we propose a “locking” strategy for the rational redesign of FAST-PETase, one of the best-performing PETase variants reported so far, to further improve its performance. The best variant, FAST-PETase DC (A171C/S193C), exhibits 1.9-fold enhanced degradation efficiency compared with FAST-PETase at 50 °C, and a 4.1 °C increase in melting temperature ( T m ). FAST-PETase DC can almost completely depolymerize untreated post-consumer PET film within 3 d at 50 °C with periodic enzyme replenishment, two-fold faster than FAST-PETase. Molecular dynamics simulations reveal that the mutation locks Helix 5 and Loop 10 through a stable disulfide bond, and reduces the flexibility of the mutation sites and their connected regions. The structural changes consequently promote the substrate binding to the enzyme, facilitate the interaction within the catalytic triad, and rigidify the overall structure of the enzyme, leading to the improved degradation efficiency and thermostability. The study underscores the “locking” strategy as an effective way to enzyme redesign, and the engineered FAST-PETase variant is a promising hydrolase for the treatment and recycling of low-to-medium crystallinity PET plastic waste.
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高影响因子
Biomolecules
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2025
·Vol. 15
·期 6
·888
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IF 4.8
Jun Yang,Binyang Deng,Pingan Liao,Siyu Lin,Liqi Zheng,Xing Yang,Fei Wang,Chao Zhai,Lixin Ma
Previous reports indicated that self-assembling amphipathic peptide S1v1 (AEAEAHAH)2significantly enhances the soluble expression, thermostability, and activity of the target proteins when fused to them. In order to obtain high-efficiency enzymes for the large-scale degradation of polyethylene terephthalate (PET), this multifunctional peptide was fused to the N- and C-terminus of FAST-PETase, a variant ofIdeonella sakaiensisPETase (IsPETase), with a PT-linker (TTVTTPQTS) harbored between the target protein and the multifunctional peptide. Consistent with previous reports, S1v1 increased the solubility of FAST-PETase slightly. Moreover, it increased the activity of FAST-PETase dramatically. The amount of terephthalic acid (TPA) and mono(2-hydroxyethyl) terephthalic acid (MHET) released from PET substrate after 24 h of digestion at 50°C by fusion enzymes bearing N- and C-terminal S1v1 tag was approximately 2.9- and 4.6-fold that of FAST-PETase, respectively. Furthermore, the optimal temperature and thermostability of the fusion proteins increased in comparison with FAST-PETase. The present study provides a novel strategy to improve the depolymerization efficiency of FAST-PETase.
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高影响因子
BIORESOURCE TECHNOLOGY
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2024
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IF 9.7
Kai Xue,Zhonghu Bai,Eric Fordour,Siqi Guo,Yuxuan Zhou,Yankun Yang,Xiuxia Liu,Ye Li,Chun-Li Liu
Biological degradation of PET plastic holds great potential for plastic recycling. However, the high costs associated with preparing free enzymes for degrading PET make it unfeasible for industrial applications. Hence, we developed various cell catalysts by surface-displaying PETase mutants and MHETase using autotransporters in E. coli and P. putida . The efficiency of surface display was enhanced through modifying the host, co-expressing molecular chaperones, and evoluting the autotransporter. In strain EC9F, PET degradation rate was boosted to 3.85 mM/d, 51-fold and 23-fold increase compared to free enzyme and initial strain ED1, respectively. The reusability of cell catalyst EC9F was demonstrated with over 38 % and 30 % of its initial activity retained after 22 cycles of BHET degradation and 3 cycles of PET degradation. The highest reported PET degradation rate of 4.95 mM/d was achieved by the dual-enzyme cascade catalytic system EC9F+EM2+R, a mixture of cell catalyst EC9F and EM2 with surfactant rhamnolipid.
研究领域:
生物化学
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环境微生物学
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合成生物学
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高影响因子
Cell Reports Physical Science
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2024
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IF 7.9
Yidi Liu,Huanliu Lin,Zhisheng Wei,Shujin Bai,Sheng Chen,Jing Wu,Zhanzhi Liu
Due to inadequate disposal and recycling of polyethylene terephthalate (PET) plastic products, there has been growing concern about the severe environmental consequences. Biocatalytic degradation of PET has emerged as a promising solution. The PET hydrolase Is PETase from Ideonella sakaiensis exhibits distinctive substrate activity at mild temperatures, but its practical application is limited by restricted catalytic efficiency. In this study, the highly flexible loops of Is PETase are targeted for engineering after a detailed structural analysis. A customized dual-fluorescence high-throughput assay is then developed to facilitate the directed evolution of these selected loops in order to enhance Is PETase’s catalytic performance, specifically at mild temperatures. Through implementation of this system, the evolved Is PETase mutant M4-Q demonstrates exceptional PET biodegradation activity at mild temperatures and advantages in degrading various aliphatic-aromatic copolyesters. These findings offer innovative insights into the enzymatic degradation of PET and other polyester plastic waste.
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高影响因子
CHINESE JOURNAL OF CHEMICAL ENGINEERING
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2023
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IF 3.8
Renwen Tian,Yan Sun
Extensive use of polyethylene terephthalate (PET) has brought about global environmental problems. A recently reported PET hydrolase (PETase) discovered from Ideonella sakaiensis showed high potential for degrading PET at moderate temperatures, but its activity and stability need further improvement for practical applications. Herein, we proposed to use α-synuclein (αS) as a fusion chaperone and created six PETase-αS fusion enzymes with linkers of different types and lengths. All the fusion enzymes exhibited improved enzymatic performance, presenting 1.5 to 2.6-fold higher activity towards bis-2(hydroxyethyl) terephthalate than PETase, as well as significantly increased stabilities. Fluorescence spectroscopy indicated that the chaperone fusion tightened the overall conformation and resulted in the opening of the substrate binding pocket, which led to the improved thermal stability and catalytic activity of the fusion enzymes. Remarkably, one of the fusion proteins, PETase-[(GS)(EK)] 10 -αS, showed 3.2 to 5.1 times higher PET degradation capability than PETase. The significantly boosted PET degradation performance was not only attributed to the enhanced enzymatic activity and stability, but also possibly due to the binding affinity of the fused αS domain for PET. These findings demonstrated that αS was an effective fusion chaperone for significantly enhancing the enzymatic performance of PETase.
研究领域:
酶学
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生物技术
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环境科学
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蛋白质工程
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Synthetic and Systems Biotechnology
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2024
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IF 4.4
Haixia Wang,Jiahong Zhu,Meng Sun,Mengjie Gu,Xiya Xie,Tongtong Ying,Zeling Zhang,Weihong Zhong
The waste pollution problem caused by polyethylene terephthalate (PET) plastics poses a huge threat to the environment and human health. As plasticizers, Phthalate esters (PAEs) are widely used in PET production and become combined pollutants with PET. Synthetic biology make it possible to construct engineered cells for microbial degradation of combined pollutants of PET and PAEs. PET hydroxylase (PETase) and monohydroxyethyl terephthalate hydroxylase (MHETase) isolated from Ideonella sakaiensis 201-F6 exhibit the capability to depolymerize PET. However, PET cannot enter cells, thus enzymatic degradation or cell surface displaying technology of PET hydrolase are the potential strategies. In this study, Pseudomonas sp. JY-Q was selected as a chassis strain, which exhibits robust stress tolerance. First, a truncated endogenous outer membrane protein cOmpA and its variant Signal (OprF)-cOmpA were selected as anchor motifs for exogenous protein to display on the cell surface. These anchor motifs were fused at the N-terminal of PET hydrolase and MHETase and transformed into Pseudomonas sp. JY-Q, the mutant strains successfully display the enzymes on cell surface, after verification by green fluorescent protein labeling and indirect immunofluorescence assay. The resultant strains also showed the catalytic activity of co-displaying PETase and MHETase for PET biodegradation. Then, the cell surface displaying PET degradation module was introduced to a JY-Q strain which genome was integrated with PAEs degrading enzymes and exhibited PAEs degradation ability. The resultant strain JY-Q-R1-R4-SFM-TPH have the ability of degradation PET and PAEs simultaneously. This study provided a promising strain resource for PET and PAEs pollution control.
研究领域:
环境科学
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微生物学
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合成生物学
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生物技术
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PubMed
高影响因子
BIORESOURCE TECHNOLOGY
·
2023
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IF 11.4
Lizhu Aer,Huiling Qin,Peng Wo,Juan Feng,Lixia Tang
The use of polyethylene terephthalate (PET) results in a significant amount of plastic waste, which poses a threat to the environment and human health. Dual-enzyme system is promising candidate for PET depolymerization. However, its production in Escherichia coli is challenging, especially for secretory expression. Herein, a novel bifunctional dual-enzyme, TfH-FPE, was constructed through fusion of FAST-PETase and TfH. TfH modifies cell membrane permeability via phospholipid degradation, thus facilitating the secretion of TfH-FPE into the medium. After systematic optimization, purified secreted TfH-FPE reached 104 ± 5.2 mg/L, which is 32.5-fold higher than that of the secreted enzyme using a signal peptide. TfH-FPE exhibits remarkable PET depolymerization capacity compared to FAST-PETase, releasing 6-fold more product than FAST-PETase and 2-fold more product than an equimolar enzyme mixture. Collectively, this study explores a novel secretory approach for efficient production of TfH-FPE and provides a valuable tool to promote PET bio-depolymerization via multi-enzyme cascades.
研究领域:
环境科学
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生物技术
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生物化学
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微生物学
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DOI
PubMed
高影响因子
BIORESOURCE TECHNOLOGY
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2024
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IF 9.7
Yu Zhou,Jiaxing Zhang,Shengping You,Wei Lin,Baoyu Zhang,Mengfan Wang,Rongxin Su,Wei Qi
Eco-friendly enzymatic-recycling has been widely utilized in tackling plastic pollution. However, the limited activity on the polyethylene terephthalate (PET) degradation product mono-hydroxyethyl terephthalate (MHET) leads to the formation of heterogeneous hydrolysis products, resulting in PET downcycling. Herein, by applying a dual-function PET hydrolase Is PETase PA with balanced PET and MHET degradation efficiency, an effective PET hydrolysis process was developed to enhance the terephthalic acid (TPA) product purity. Firstly, the impact of pH on the catalytic activity of Is PETase PA revealed that the pH reduction caused by TPA generation hindered the complete conversion of MHET to TPA. Further investigation of the catalytic mechanism showed that the pH-induced protonation of H208 in the catalytic triad destabilized the interaction between Is PETase PA and MHET. Thus, by introducing pH regulation strategy on the bifunctional Is PETase PA , the single-enzyme process could achieve high-purity TPA recovery (>99 %). Overall, this work ensured the high-quality PET enzymatic-recycling for effectively addressing plastic pollution.
研究领域:
环境科学
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生物技术
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材料科学
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化学工程
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高影响因子
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