Comparative performance of PETase as a function of reaction conditions, substrate properties, and product accumulation

Erika Erickson, Thomas Jack Shakespeare, Felicia Bratti, Bonnie Buss, Rosie Graham, McKenzie Hawkins, Gerhard Koenig, William E. Michener, Joel Miscall, Kelsey Ramirez, Nicholas A. Rorrer, Michael Zahn, Andrew Pickford, John McGeehan*, Gregg T. Beckham*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

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There is keen interest to develop new technologies to recycle the plastic poly(ethylene terephthalate) (PET). To this end, the use of PET-hydrolyzing enzymes has shown promise for PET deconstruction to its monomers, terephthalate (TPA) and ethylene glycol (EG). Here, we compare the Ideonella sakaiensis PETase wild-type enzyme to a previously reported improved variant (W159H/S238F). We compare the thermostability of each enzyme and describe a 1.45 Å resolution structure of the mutant, highlighting changes in the substrate binding cleft compared to the wild-type enzyme. Subsequently, the performance of the wild-type and variant enzyme was compared as a function of temperature, substrate morphology, and reaction mixture composition. These studies show that reaction temperature has the strongest influence on performance between the two enzymes. We also show that both enzymes achieve higher levels of PET conversion for substrates with moderate crystallinity relative to amorphous substrates. Finally, we assess the impact of product accumulation on reaction progress for the hydrolysis of both PET and bis(2-hydroxyethyl) terephthalate (BHET). Each enzyme displays different inhibition profiles to mono(2-hydroxyethyl) terephthalate (MHET) and TPA, while both are sensitive to inhibition by EG. Overall, this study highlights the importance of reaction conditions, substrate selection, and product accumulation for catalytic performance of PET-hydrolyzing enzymes, which have implications for enzyme screening in the development of enzyme-based polyester recycling.
Original languageEnglish
Article numbere202101932
Number of pages11
Issue number1
Early online date5 Nov 2021
Publication statusPublished - 10 Jan 2022


  • Chemical recycling
  • interfacial biocatalysis
  • IsPETase
  • kinetics
  • Michaelis-Menten
  • PET hydrolase
  • UKRI
  • BB/P011918/1


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