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Methods in Enzymology
Volume 648, 2021, Pages 187-205

Ideonella sakaiensis, PETase, and MHETase: From identification of microbial PET degradation to enzyme characterization

Shosuke Yoshidaa, Kazumi Hiragab, Ikuo Taniguchic, Kohei Odab

Institute for Research Initiatives, Nara Institute of Science and Technology & Division of Biological Science, Nara Institute of Science and Technology, Ikoma, Nara, Japan.

Abstract

Few reports have described the biological degradation or utilization of poly(ethylene terephthalate) (PET) to support microbial growth. We screened environmental samples from a PET bottle recycling site and identified the microbial consortium no. 46, which degraded amorphous PET at ambient temperature; thereafter, we isolated the resident Ideonella sakaiensis 201-F6 strain responsible for the degradation. We further identified two hydrolytic enzymes from I. sakaiensis, PET hydrolase (PETase) and mono(2-hydroxyethyl) terephthalate hydrolase (MHETase), which synergistically converted PET into its monomeric building blocks. Here, we provide original methods of microbial screening and isolation of PET degrading microbe(s). These novel approaches can be adapted for exploring microorganisms that degrade PET and other plastics. Furthermore, our enzyme assay protocols to characterize PETase and MHETase can be applied to evaluate new enzymes that target PET and its hydrolysates.

Keywords: Poly(ethylene terephthalate), Microbial consortium, Ideonella sakaiensis, PET hydrolase, Mono(2-hydroxyethyl) terephthalic acid hydrolase, PET hydrolytic enzyme (PHE).

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