Engineering dual-functional and thermophilic BMHETase for efficient degradation of polyethylene terephthalate

Polyethylene terephthalate (PET) biodegradation is hindered by the intermediates bis (2-hydroxyethyl) terephthalate (BHET) and mono (2-hydroxyethyl) terephthalate (MHET). BMHETase, a thermophilic hydrolase identified from the UniParc database, exhibits degradation activity towards both BHET and MHET. BMHETase showed higher activity on BHET than LCCICCG and FASTPETase at temperatures ranging from 50 to 70℃. To enhance its activity in degrading MHET, BMHETase was engineered to mimic Ideonella sakaiensis MHETase. The resulting 6-point mutant's activities on MHET and BHET were 8 and 2 times those of the WT, with both optimal temperatures increased by 5℃. This enhancement may be attributed to the BMHETase^6M's intensified binding ability with MHET and enlarged binding pocket. When combined with LCCICCG, BMHETase^6M achieved complete degradation of MHET in PET films to terephthalic acid, indicating broad application potential. These findings suggest that BMHETase^6M holds promise as a candidate for enhancing PET biodegradation efficiency and plastic waste management.