Functional tailoring of a PET hydrolytic enzyme expressed in Pichia pastoris

Li, Xian; Shi, Beilei; Huang, Jian‐Wen; Zeng, Ziyin; Yang, Yu; Min, Jian; Chen, Chun‐Chi; Guo, Rey‐Ting

doi:10.1186/s40643-023-00648-1

Cited by 8 publications

(8 citation statements)

References 41 publications

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“…5b, iii) (Chen et al 2021a, b). F235L alteration further elevates the activity by 60% at 60 °C, and introducing DS3 by R230C-S284C mutation enhances the enzyme activity at 70 °C by more than 2.5-fold (Li et al 2023). Notably, CtPL-DM showed no activity when expressed in Pichia pastoris.…”

Section: Type II Enzyme Engineeringmentioning

confidence: 98%

“…This could be an obstacle for PET-degrading enzymes, as measuring PET hydrolytic products by highperformance liquid chromatography (HPLC) analysis is the standard method to probe the efficacy of these enzymes. Shi et al ( 2023) developed a novel PET analogous substrate, bis (2-hydroxyethyl) 2-hydroxyterephthalate (Shi et al 2023). The enzyme activity can be quickly determined by measuring the fluorescence emitted by the hydrolyzed product.…”

Section: Type II Enzyme Engineeringmentioning

confidence: 99%

“…This has been suspected due to steric hindrance mediated by the N-glycosylation on N181 that constrains the flexibility of the + 1 subsite-forming Trp. N181A mutation that reduced glycosylation degree completely restored the enzyme activity (Li et al 2023). These results have important application implications as P. pastoris is a workhorse in enzyme production industries and could be a candidate strain for large-scale production of PET decomposing enzymes.…”

Section: Type II Enzyme Engineeringmentioning

confidence: 99%

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Natural and engineered enzymes for polyester degradation: a review

Guo,

Li,

Yang

et al. 2024

Environ Chem Lett

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Plastic pollution is becoming a major health issue due to the recent discovery of microplastics and nanoplastics in living organisms and the environment, calling for advanced technologies to remove plastic waste. Here we review enzymes that degrade plastics with focus on plastic properties, protein engineering and polymers such as poly(ethylene terephthalate), poly(butylene adipate-co-terephthalate), poly(lactic acid), polyamide and polyurethane. The mechanism of action of natural and engineered enzymes has been probed by experimental and computation approaches. The performance of polyester-degrading enzymes has been improved via directed evolution, structure-guided rational design and machine learning-aided strategies. The improved enzymes display higher stability at elevated temperatures, and tailored substrate-binding sites.

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Section: Type II Enzyme Engineeringmentioning

confidence: 98%

Section: Type II Enzyme Engineeringmentioning

confidence: 99%

Section: Type II Enzyme Engineeringmentioning

confidence: 99%

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Natural and engineered enzymes for polyester degradation: a review

Guo,

Li,

Yang

et al. 2024

Environ Chem Lett

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“…Type I enzymes share His164 and Phe243 residues in subsite II, except PHL7, where Phe is replaced with Leu (Sonnendecker et al 2022 ). Type IIb enzymes have Trp and Ser/Thr at the corresponding positions, whereas type IIa enzymes have Trp and Phe/Tyr at the same position, except for PET2, where it is Trp and Trp (Meilleur et al 2009 ; Danso et al 2018 ; Nakamura et al 2021 ), and Ct PL, where it is His and Phe (Chen et al 2021 ; Li et al 2023 ). His, Phe, Trp, and Tyr are aromatic amino acids and can replace each other.…”

Section: Pet Hydrolasesmentioning

confidence: 99%

Engineered polyethylene terephthalate hydrolases: perspectives and limits

Kawai,

Iizuka,

Kawabata

2024

Appl Microbiol Biotechnol

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Polyethylene terephthalate (PET) is a major component of plastic waste. Enzymatic PET hydrolysis is the most ecofriendly recycling technology. The biorecycling of PET waste requires the complete depolymerization of PET to terephthalate and ethylene glycol. The history of enzymatic PET depolymerization has revealed two critical issues for the industrial depolymerization of PET: industrially available PET hydrolases and pretreatment of PET waste to make it susceptible to full enzymatic hydrolysis. As none of the wild-type enzymes can satisfy the requirements for industrialization, various mutational improvements have been performed, through classical technology to state-of-the-art computational/machine-learning technology. Recent engineering studies on PET hydrolases have brought a new insight that flexibility of the substrate-binding groove may improve the efficiency of PET hydrolysis while maintaining sufficient thermostability, although the previous studies focused only on enzymatic thermostability above the glass transition temperature of PET. Industrial biorecycling of PET waste is scheduled to be implemented, using micronized amorphous PET. Next stage must be the development of PET hydrolases that can efficiently degrade crystalline parts of PET and expansion of target PET materials, not only bottles but also textiles, packages, and microplastics. This review discusses the current status of PET hydrolases, their potential applications, and their profespectal goals. Key points • PET hydrolases must be thermophilic, but their operation must be below 70 °C • Classical and state-of-the-art engineering approaches are useful for PET hydrolases • Enzyme activity on crystalline PET is most expected for future PET biorecycling Graphical Abstract

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“…The study reported that N ‐glycosylation at N181 of Caldimonas taiwanensis PETase resulted in restrained conformation change of Trp present in the binding domain, which is essential for the activity of the PETase. [ 184 ]…”

Section: Bioremediation Of Pnps: Plastic Degrading Proteins and Their...mentioning

confidence: 99%

Mechanistic Insights into Cellular and Molecular Basis of Protein‐Nanoplastic Interactions

Naidu,

Nagar,

Poluri

2023

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Plastic waste is ubiquitously present across the world, and its nano/sub‐micron analogues (plastic nanoparticles, PNPs), raise severe environmental concerns affecting organisms’ health. Considering the direct and indirect toxic implications of PNPs, their biological impacts are actively being studied; lately, with special emphasis on cellular and molecular mechanistic intricacies. Combinatorial OMICS studies identified proteins as major regulators of PNP mediated cellular toxicity via activation of oxidative enzymes and generation of ROS. Alteration of protein function by PNPs results in DNA damage, organellar dysfunction, and autophagy, thus resulting in inflammation/cell death. The molecular mechanistic basis of these cellular toxic endeavors is fine‐tuned at the level of structural alterations in proteins of physiological relevance. Detailed biophysical studies on such protein‐PNP interactions evidenced prominent modifications in their structural architecture and conformational energy landscape. Another essential aspect of the protein‐PNP interactions includes bioenzymatic plastic degradation perspective, as the interactive units of plastics are essentially nano‐sized. Combining all these attributes of protein‐PNP interactions, the current review comprehensively documented the contemporary understanding of the concerned interactions in the light of cellular, molecular, kinetic/thermodynamic details. Additionally, the applicatory, economical facet of these interactions, PNP biogeochemical cycle and enzymatic advances pertaining to plastic degradation has also been discussed.

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Functional tailoring of a PET hydrolytic enzyme expressed in Pichia pastoris

Cited by 8 publications

References 41 publications

Natural and engineered enzymes for polyester degradation: a review

Natural and engineered enzymes for polyester degradation: a review

Engineered polyethylene terephthalate hydrolases: perspectives and limits

Mechanistic Insights into Cellular and Molecular Basis of Protein‐Nanoplastic Interactions

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