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Directed evolution of an efficient and thermostable PET depolymerase

DOI: 10.1038/s41929-022-00821-3 DOI Help

Authors: Elizabeth L. Bell (University of Manchester) , Ross Smithson (University of Manchester) , Siobhan Kilbride (University of Manchester) , Jake Foster (University of Manchester) , Florence J. Hardy (University of Manchester) , Saranarayanan Ramachandran (University of Manchester) , Aleksander A. Tedstone (University of Manchester) , Sarah J. Haigh (University of Manchester) , Arthur A. Garforth (University of Manchester) , Philip J. R. Day (University of Manchester) , Colin Levy (University of Manchester) , Michael P. Shaver (University of Manchester) , Anthony P. Green (University of Manchester)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Nature Catalysis , VOL 3

State: Published (Approved)
Published: August 2022
Diamond Proposal Number(s): 12788 , 17773

Abstract: The recent discovery of IsPETase, a hydrolytic enzyme that can deconstruct poly(ethylene terephthalate) (PET), has sparked great interest in biocatalytic approaches to recycle plastics. Realization of commercial use will require the development of robust engineered enzymes that meet the demands of industrial processes. Although rationally engineered PETases have been described, enzymes that have been experimentally optimized via directed evolution have not previously been reported. Here, we describe an automated, high-throughput directed evolution platform for engineering polymer degrading enzymes. Applying catalytic activity at elevated temperatures as a primary selection pressure, a thermostable IsPETase variant (HotPETase, Tm = 82.5 °C) was engineered that can operate at the glass transition temperature of PET. HotPETase can depolymerize semicrystalline PET more rapidly than previously reported PETases and can selectively deconstruct the PET component of a laminated multimaterial. Structural analysis of HotPETase reveals interesting features that have emerged to improve thermotolerance and catalytic performance. Our study establishes laboratory evolution as a platform for engineering useful plastic degrading enzymes.

Journal Keywords: Biocatalysis; Environmental biotechnology; Hydrolases; Protein design

Diamond Keywords: Plastics; Enzymes; Biodegradation

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

Instruments: I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography

Added On: 16/08/2022 20:38

Discipline Tags:

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

Technical Tags:

Diffraction Macromolecular Crystallography (MX)