(515c) Efficient Biodegradation of Poly(ethylene Terephthalate) with Leaf-Branch Compost Cutinase

Authors: 
Liu, N., University of Massachusetts Lowell
Xie, D., University of Massachusetts-Lowell
Patel, A., University of Massachusetts Lowell
Wong, H. W., University of Massachusetts Lowell
Sobkowicz, M. J., University of Massachusetts Lowell
Poly(ethylene terephthalate) (PET) is one of the most widely used polyester plastics in the world, but is extremely difficult to be hydrolyzed. Accumulation of the PET in the environment at a staggering rate as discarded packaging and textiles pose a global environmental problem. Cutinases are polyester hydrolases that show a remarkable capability to PET to its monomeric units. This revelation has stimulated research aimed at developing sustainable and green cutinase-catalyzed PET biorecycling methods. The gene encoding a cutinase homolog, LCC, was cloned from a fosmid library of a leaf-branch compost metagenome by functional screening using tributyrin agar plates. LCC is particularly suited toward these ends given its relatively high PET hydrolysis activity and thermostability. With the assistance of the pelB leader sequence, more than 36% of the recombinant LCC produced by Escherichia coli BL21(DE3) cells was excreted into the culture medium. To improve the biodegradation efficiency, recycled PET (RPET) was cut into small pieces via grinding to decrease PET particle size and increase surface area. Different particle size of RPET were examined. As a result, the degradation rate of the LCC in the presence of smaller RPET particle size (with 60 mesh screen) was significantly improved, with a productivity of 1.2 milligrams of terephthalic acid (TPA) per milliliter over 48 hours (with enzyme concentration of 0.8 micrograms per milligram of PET). Moreover, the substrate-binding modules from a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (PBM) was fused with the linker (2.3 kDa) from a cellobiohydrolase from Trichoderma reesei to the C-terminus of LCC to improve the PET adsorption. Although PBM (6.3 kDa) binding modules have a hydrophobic nature, it was possible to express the proteins in E. coli. Our result exhibited that adsorption to PET by the LCC-PBM fusion enzyme was strikingly enhanced as compared with LCC. Though the PET hydrolysis performance of any LCC variants was much higher than that of the bacterial enzyme Is-PETase from Ideonella sakaiensis 201-F6, the degradation efficiency of LCC-PBM into TPA was not as high as LCC. This study provides us a great potential to achieve enzymatically depolymerization of PET waste contributing towards the concept of a circular PET economy.