Microplastic (MP) pollution is a growing environmental concern, with wastewater treatment plants (WWTPs) serving as a critical yet insufficient barrier. Although WWTPs can remove up to 99% of MPs from influent streams, a large fraction of the sequestered MPs is concentrated into sewage sludge and subsequently redistributed to soil and waterways via biosolids. Here, we report a biosynthetic approach to address MP redistribution by coupling microplastic removal with catalytic depolymerization, using enzyme-functionalized bacterial nanocellulose (BNC) filtration materials. As a proof-of-concept, we fused a cellulose-binding domain (CBD) to the thermostable PET hydrolase HotPETase, creating a bifunctional protein, CBD-PETase, that binds to a BNC scaffold and degrades PET microplastic (PET) captured within the scaffold. We showed that BNC-CBD-PETase composites sequestered irregularly shaped PET within hierarchical pore networks and generated PET degradation products proportionally to enzyme loading. We further established a co-culture strategy, in which Saccharomyces cerevisiae was engineered to secrete CBD-PETase during BNC scaffold biosynthesis by Komagataeibacter xylinus, which enabled one-pot fabrication of an active BNC-CBD-PETase composite without separate protein purification. Notably, co-culture-derived BNC-CBD-PETase composites retained catalytic activity for at least 24 weeks under dry, ambient storage. This platform provides a route towards a scalable and renewable filtration material with the potential for integration across multiple treatment stages within WWTPs.
Pitt, B. N., Guillen, D., Obermeyer, A. C.
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