Poly(ethylene terephthalate) (PET) is the most abundant polyester plastic and a major contributor to plastic pollution. IsPETase, from the PET-assimilating bacterium Ideonella sakaiensis, is a unique PET-hydrolytic enzyme that shares high sequence identity to canonical cutinases, but shows substrate preference towards PET and exhibits higher PET-hydrolytic activity at ambient temperature. Structural analyses suggest that IsPETase harbours a substrate-binding residue, W185, with a wobbling conformation and a highly flexible W185-locating β6-β7 loop. Here, we show that these features result from the presence of S214 and I218 in IsPETase, whose equivalents are strictly His and Phe, respectively, in all other homologous enzymes. We found that mutating His/Phe residues to Ser/Ile could enhance the PET-hydrolytic activity of several IsPETase-like enzymes. In conclusion, the Ser/Ile mutations should provide an important strategy to improve the activity of potential PET-hydrolytic enzymes with properties that may be useful for various applications.
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Source data are provided with this paper. The atomic coordinates and structure factors of the reported structures have been deposited in the Protein Data Bank under accession codes 7CY0 for IsPETase S214H and 7CWQ for BurPL. The web links of previously reported protein structures that were utilized for analyses and comparisons in this study are provided in the figure legends. Data and other findings of this study are available from the corresponding author on reasonable request.
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This work was supported by the National Key Research and Development Program of China (2019YFA0706900, R.-T.G.), the National Natural Science Foundation of China (31870790, R.-T.G.; 31971205, C.-C.C.; 31800662, X.H.; 32000899, L.Z.) and the Natural Science Foundation Innovative Group Project of Hubei Province (2020CFA011, R.-T.G.). We thank X-ray crystallography facility of State Key Laboratory of Biocatalysis and Enzyme Engineering and National Synchrotron Radiation Research Center (NSRRC) for access to beamlines TPS-05A and TLS-15A1, which contributed to the synchrotron data collection.
The authors declare no competing interests.
Peer review information Nature Catalysis thanks Kohei Oda, César A. Ramírez-Sarmiento and Mitch H. Weiland for their contribution to the peer review of this work.
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Chen, CC., Han, X., Li, X. et al. General features to enhance enzymatic activity of poly(ethylene terephthalate) hydrolysis. Nat Catal 4, 425–430 (2021). https://doi.org/10.1038/s41929-021-00616-y