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Engineering >> 2018, Volume 4, Issue 6 doi: 10.1016/j.eng.2018.09.007

Enhanced Poly(ethylene terephthalate) Hydrolase Activity by Protein Engineering

a Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China

b SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering, Tianjin University, Tianjin 300072, China

Received: 2018-03-02 Revised: 2018-05-18 Accepted: 2018-09-14 Available online: 2018-09-21

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Abstract

Poly(ethylene terephthalate) hydrolase (PETase) from Ideonella sakaiensis exhibits a strong ability to degrade poly(ethylene terephthalate) (PET) at room temperature, and is thus regarded as a potential tool to solve the issue of polyester plastic pollution. Therefore, we explored the interaction between PETase and the substrate (a dimer of the PET monomer ethylene terephthalate, 2PET), using a model of PETase and its substrate. In this study, we focused on six key residues around the substrate-binding groove in order to create novel high-efficiency PETase mutants through protein engineering. These PETase mutants were designed and tested. The enzymatic activities of the R61A, L88F, and I179F mutants, which were obtained with a rapid cell-free screening system, exhibited 1.4 fold, 2.1 fold, and 2.5 fold increases, respectively, in comparison with wild-type PETase. The I179F mutant showed the highest activity, with the degradation rate of a PET film reaching 22.5 mg per lmol·L–1 PETase per day. Thus, this study has created enhanced artificial PETase enzymes through the rational protein engineering of key hydrophobic sites, and has further illustrated the potential of biodegradable plastics.

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