Purification of high-temperature resistant polyethylene terephthalate (PET) hydrolase by simple heating protocol

In this study, a novel high-purity polyethylene terephthalate (PET) hydrolase purification method was suggested, which was based on a simple heating protocol. To test the applicability of the method, the heating purification performances of different types of thermo-resistant PET hydrolases, including ThcCut1, ThcCut2, TfCut2, TfCut2-G62A, TfCut2-G62A/F209A, and LCC-F243I/D238C/S283C/Y127G, from crude extract were conducted. During the process, the effect of protectants and treatment duration on proteins and activity recoveries were tested. Results indicated that the specific activities of PET hydrolases purified by heating were generally as competitive as the ones purified by conventional nickel column methods, with higher protein recovery and enzyme activity recovery. The practical application of the heating purified PET hydrolases was further evaluated. Except for ThcCut2, other PET hydrolases exhibited better degradation performances of PET nanoparticles. The novel heating purification method for PET hydrolases showed good applicability, which can be extended to purify other thermo-resistant enzymes.