Complete bio-degradation of poly(butylene adipate-co-terephthalate) via engineered cutinases


  Poly(butylene adipate-co-terephthalate) (PBAT) is a synthetic polymer made of adipic acid, butylene glycol, and terephthalic acid (see the figure below). Because of its excellent ductility, thermal stability, and plasticity, PBAT is widely used in agricultural industries, textile manufacturing, and food packaging. Despite being named as a “biodegradable plastic”, the complete decomposition of PBAT in nature proceeds at a very low rate, which has brought severe concerns. This is exemplified by the broadly utilized mulching films, which are left in the farmland and shattered into smaller fragments of plastics instead of being completely decomposed. These fragments are mixed with soil to result in soil agglomeration and reduce land productivity. The mechanistic study of PBAT degradation is impeded due to the complicated structure of PBAT. Moreover, PBAT mulch film can be cross-linked when exposed to UV light, making PBAT more difficult to degrade. Therefore, we seek a method that can rapidly and efficiently degrade polymer PBAT and elucidate its degradation mechanism.

  Recently, our team discovered that a cutinase from Thermobifida fusca (TfCut) can break down PBAT mulch film into large fragments, and small particles until they disappear completely within two days. Further separation and identification of intermediates showed three main intermediate products, namely BTa, ABTa, TaBTa (Ta=TPA), and the final product TPA. Interestingly, there are many possibilities for the products of PBAT degradation, however, only these four products appear in the process of TfCut-mediated degradation of PBAT. It is worth noting that all four products have TPA as the terminus, suggesting a unique degradation mechanism of TfCut in the degradation of PBAT.

  We later traced the changes in four products and found that after reaching their highest values in about 8 hours, ABTa and TaBTa gradually decreased until non-detectable. After 48 hours, the final products were TPA and BTa. Moreover, they previously applied the strategy by replacing the His/Phe due (H224-F228) with the Ser/Ile due (S224-I228) in TfCut, and the capacity of degrading PET was significantly enhanced (Chen et al., Nature Catalysis, 2021, 4, p. 425). Therefore, after applying this strategy to the degradation of PBAT, the result shows that only TPA remained after 48 hours of the engineered TfCut (referred to as TfCut-DM) in the degradation reaction, which will be easier to recycle the TPA after the degradation of PBAT in the future.

  We also obtained the complex structures of wild-type TfCut and TfCut-DM with the substrate analog MHET. Both the wild-type TfCut and reported LCC-ICCG to contain the His/Phe due that forms a barrier at the entrance of their active site. In contrast, TfCut-DM and IsPETase have the Ser/Ile due and leads to a more open and flat TPA-binding pocket, which produces less hindrance for the entry of bulkier substrates.

  Above all, our team drew a schematic diagram of the TfCut-mediated degradation of PBAT. TfCut first performs the endo-hydrolase function with its catalytic triad. As a result, intermediates such as BTa, ABTa, and TaBTa, have the TPA as the terminal. The end products after degradation by wild-type TfCut are BTa and TPA, while they become TPA in the TfCut-DM mediate degradation, which will be more conducive to the recovery and recycling of PBAT.

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