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Conformational Selection in Biocatalytic Plastic Degradation by PETase
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Fibre- and Polymer Technology, Coating Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Fibre- and Polymer Technology, Coating Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-8195-1099
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Fibre- and Polymer Technology, Coating Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-4708-9861
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Fibre- and Polymer Technology, Coating Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-6711-4972
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2022 (English)In: ACS Catalysis, E-ISSN 2155-5435, Vol. 12, no 6, p. 3397-3409Article in journal (Refereed) Published
Abstract [en]

Due to the steric effects imposed by bulky polymers, the formation of catalytically competent enzyme and substrate conformations is critical in the biodegradation of plastics. In poly(ethylene terephthalate) (PET), the backbone adopts different conformations, gauche and trans, coexisting to different extents in amorphous and crystalline regions. However, which conformation is susceptible to biodegradation and the extent of enzyme and substrate conformational changes required for expedient catalysis remain poorly understood. To overcome this obstacle, we utilized molecular dynamics simulations, docking, and enzyme engineering in concert with high-resolution microscopy imaging and solid-state nuclear magnetic resonance (NMR) to demonstrate the importance of conformational selection in biocatalytic plastic hydrolysis. Our results demonstrate how single-amino acid substitutions in Ideonella sakaiensis PETase can alter its conformational landscape, significantly affecting the relative abundance of productive ground-state structures ready to bind discrete substrate conformers. We experimentally show how an enzyme binds to plastic and provide a model for key residues involved in the recognition of gauche and trans conformations supported by in silico simulations. We demonstrate how enzyme engineering can be used to create a trans-selective variant, resulting in higher activity when combined with an all-trans PET-derived oligomeric substrate, stemming from both increased accessibility and conformational preference. Our work cements the importance of matching enzyme and substrate conformations in plastic hydrolysis, and we show that also the noncanonical trans conformation in PET is conducive for degradation. Understanding the contribution of enzyme and substrate conformations to biocatalytic plastic degradation could facilitate the generation of designer enzymes with increased performance.

Place, publisher, year, edition, pages
American Chemical Society (ACS) , 2022. Vol. 12, no 6, p. 3397-3409
Keywords [en]
PET, plastic biodegradation, conformational selection, gauche/trans, enzyme engineering
National Category
Biochemistry Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-311888DOI: 10.1021/acscatal.1c05548ISI: 000778789200014Scopus ID: 2-s2.0-85126103731OAI: oai:DiVA.org:kth-311888DiVA, id: diva2:1656626
Note

QC 20220524

Available from: 2022-05-06 Created: 2022-05-06 Last updated: 2025-02-20Bibliographically approved
In thesis
1. Chemoenzymatic Synthesis and Degradation of Plastics
Open this publication in new window or tab >>Chemoenzymatic Synthesis and Degradation of Plastics
2024 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The development of a carbon-based bioeconomy for synthesis and degradation of polymers has gained importance over the years. Research efforts have been made to develop green routes to produce bio-based material from biomass as well as environmentally friendly ways to synthesize and degrade polymers. Enzymes are biocatalysts that are capable of performing reactions beyond their intended purpose. The work presented in this thesis focused on using biocatalysts for novel reactions to produce bio-plastics as well as degrade synthetic polymers. In Paper I, a decarboxylase was used to perform the fixation of CO2 under mild conditions to produce the platform chemical 2,5-furandicarboxylic acid (FDCA). In Paper II, a closed-loop approach for the production of bio-based polyesters and their enzymatic degradation was investigated. Moreover, the difference of catalytic activity towards different polymer conformations was noted and further investigated in Paper III. Here, the conformational landscape to match enzyme to substrate was explored. The model substrate for this project was post-consumer PET bottles since is one of the most used polymer worldwide. The substrate conformation affected the catalytic activity of the enzymes significantly hence, in Paper IV, the physical and chemical characteristics of various PET-based substrates was investigated to better understand the factors that will yield a high reaction efficiency for polymer depolymerization. Finally, the results obtained so far were used in Paper V to show that plastic degrading enzymes can be used for microplastic degradation in human blood as a proof-of-concept. In summary, the work in this thesis showed the potential of using enzymes as catalysts for the production of platform chemicals through CO2 fixation and for polymer degradation initiating an attractive path to close the loop in a bio-economy for polymer materials.

Abstract [sv]

Utvecklingen av en kolbaserad bioekonomi för syntes och återvinning av polymerer har blivit allt viktigare under åren. Forskningsinsatser har genomförts för att utveckla grönare alternativ för att producera biobaserade material samt miljövänliga sätt att syntetisera och bryta ner polymerer. Enzymer är biokatalysatorer som kan utföra reaktioner utöver deras avsedda syfte. Arbetet som presenteras i denna avhandling fokuserade på att använda biokatalysatorer för nya reaktioner för att producera bioplaster och bryta ner syntetiska polymerer. I Artikel I användes ett dekarboxylas för att fixera CO2 under milda förhållanden för att producera plattformskemikalien 2,5-furandikarboxylsyra (FDCA). I Artikel II undersöktes en sluten loop-metod för produktion av biobaserade polyester och deras enzymatiska nedbrytning. Dessutom noterades skillnaden i katalytisk aktivitet mot olika polymerkonformationer och undersöktes vidare i Artikel III. Här studerades det konformationella landskapet för att matcha enzym mot substrat. Modellsubstratet för detta projekt var PET-flaskor eftersom det är den mest använda polymeren världen över. Substratkonformationen påverkade enzymernas katalytiska aktivitet signifikant, varför de fysikaliska och kemiska egenskaperna hos olika PET-baserade substrat undersöktes i Artikel IV för att bättre förstå de faktorer som ger en hög reaktionseffektivitet för depolymerisering. Slutligen användes de hittills erhållna resultaten i Artikel V för att visa att plastnedbrytande enzymer kan användas för nedbrytning av mikroplast i humanblod som en proof-of-concept. Sammanfattningsvis visade arbetet i denna avhandling potentialen att använda enzymer som katalysatorer för produktion av plattformskemikalier genom CO2 -fixering och för polymernedbrytning vilket initierar en attraktiv väg för att sluta kretsloppet i en bioekonomi för polymera material. 

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2024. p. 81
Series
TRITA-CBH-FOU ; 2024:19
Keywords
Bio-based materials, Bioeconomy, Biomass, CO2, Conformational selection, Enzyme- engineering, Microwave pre-treatment, Plastic degradation, Polyester, Recycling., Biobaserade material, Bioekonomi, Biomassa, Konformationsselektion, Enzymingenejörskonst, Mikrovågsförbehandling, Plastnedbrytning, Polyester, Återvinning.
National Category
Biocatalysis and Enzyme Technology
Research subject
Fibre and Polymer Science
Identifiers
urn:nbn:se:kth:diva-346252 (URN)978-91-8040-920-9 (ISBN)
Public defence
2024-06-05, F3, Lindstedtsvägen 26, https://kth-se.zoom.us/j/62551057307, Stockholm, 10:00 (English)
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Supervisors
Note

QC 20240508

Available from: 2024-05-08 Created: 2024-05-08 Last updated: 2024-05-29Bibliographically approved

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Guo, BoyangVanga, Sudarsana ReddyLopez-Lorenzo, XimenaSaenz-Mendez, PatriciaFang, YuanYe, XinchenSchriever, KarenBäckström, EvaBiundo, AntoninoFuro, IstvanHakkarainen, MinnaSyrén, Per-Olof

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Guo, BoyangVanga, Sudarsana ReddyLopez-Lorenzo, XimenaSaenz-Mendez, PatriciaEricsson, Sara RönnbladFang, YuanYe, XinchenSchriever, KarenBäckström, EvaBiundo, AntoninoZubarev, Roman A.Furo, IstvanHakkarainen, MinnaSyrén, Per-Olof
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Coating TechnologyScience for Life Laboratory, SciLifeLabApplied Physical ChemistryPolymeric MaterialsPolymer TechnologyWallenberg Wood Science Center
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