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Sourcing thermotolerant poly(ethylene terephthalate) hydrolase scaffolds from natural diversity

DOI: 10.1038/s41467-022-35237-x DOI Help

Authors: Erika Erickson (National Renewable Energy Laboratory; BOTTLE Consortium) , Japheth E. Gado (National Renewable Energy Laboratory; BOTTLE Consortium) , Luisana Avilán (University of Portsmouth) , Felicia Bratti (National Renewable Energy Laboratory; BOTTLE Consortium) , Richard K. Brizendine (National Renewable Energy Laboratory; BOTTLE Consortium) , Paul A. Cox (University of Portsmouth) , Raj Gill (University of Portsmouth) , Rosie Graham (University of Portsmouth) , Dong-Jin Kim (BOTTLE Consortium; Montana State University) , Gerhard König (University of Portsmouth) , William E. Michener (National Renewable Energy Laboratory; BOTTLE Consortium) , Saroj Poudel (Montana State University) , Kelsey J. Ramirez (National Renewable Energy Laboratory; BOTTLE Consortium) , Thomas J. Shakespeare (University of Portsmouth) , Michael Zahn (University of Portsmouth) , Eric S. Boyd (Montana State University) , Christina M. Payne (National Science Foundation) , Jennifer L. Dubois (BOTTLE Consortium; Montana State University) , Andrew R. Pickford (BOTTLE Consortium; University of Portsmouth) , Gregg T. Beckham (National Renewable Energy Laboratory; BOTTLE Consortium) , John E. Mcgeehan (BOTTLE Consortium; University of Portsmouth; World Plastics Association)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Nature Communications , VOL 13

State: Published (Approved)
Published: December 2022
Diamond Proposal Number(s): 23269

Open Access Open Access

Abstract: Enzymatic deconstruction of poly(ethylene terephthalate) (PET) is under intense investigation, given the ability of hydrolase enzymes to depolymerize PET to its constituent monomers near the polymer glass transition temperature. To date, reported PET hydrolases have been sourced from a relatively narrow sequence space. Here, we identify additional PET-active biocatalysts from natural diversity by using bioinformatics and machine learning to mine 74 putative thermotolerant PET hydrolases. We successfully express, purify, and assay 51 enzymes from seven distinct phylogenetic groups; observing PET hydrolysis activity on amorphous PET film from 37 enzymes in reactions spanning pH from 4.5–9.0 and temperatures from 30–70 °C. We conduct PET hydrolysis time-course reactions with the best-performing enzymes, where we observe differences in substrate selectivity as function of PET morphology. We employed X-ray crystallography and AlphaFold to examine the enzyme architectures of all 74 candidates, revealing protein folds and accessory domains not previously associated with PET deconstruction. Overall, this study expands the number and diversity of thermotolerant scaffolds for enzymatic PET deconstruction.

Journal Keywords: Biocatalysis; Hydrolases

Diamond Keywords: Enzymes; Plastics; Biodegradation

Subject Areas: Chemistry, Biology and Bio-materials, Environment

Instruments: I03-Macromolecular Crystallography

Added On: 23/12/2022 08:32


Discipline Tags:

Earth Sciences & Environment Biotechnology Biochemistry Catalysis Chemistry Structural biology Engineering & Technology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)