Abstract
The escalating plastics crisis, exacerbated by the accumulation of nonbiodegradable polyesters in the environment, has necessitated the exploration of sustainable waste management solutions such as enzymatic hydrolysis in industrial recycling. So far, the focus of these efforts has been on cutinase-related polyethylene terephthalate (PET) degrading carboxylesterases, or PETases. In this work, we report the discovery and initial activity screen of previously uncharacterized, thermostable enzymes with polyesterase activity through comprehensive phylogenetic and sequence analysis of a bacterial family of esterases, Lipase Family 1.5. These enzymes are related to the previously identified polybutylene succinate co-terephthalate (PBAT) degrading carboxylesterases Cl_EstA and Cl_EstB from Clostridium botulinum and PfL1 from Pelosinus fermentans. Originating from thermophilic bacteria, we show that these enzymes can be expressed heterologously in Escherichia coli and degrade the polyesters PBAT and polybutylene succinate co-butylene adipate (PBSA), though they exhibit limited activity against PET. Notably, our results show that these enzymes are more effective at degrading the fully aliphatic polyester PBSA compared to the aliphatic-aromatic co-polyester PBAT, with three members of this enzyme family achieving complete solubilization of 5 mg/mL milled PBSA within 2 days at a low enzyme concentration (100 nM). This study highlights the substantial opportunity to find novel enzymes from nature that possess the required thermal stability for industrial applications, potentially reducing the need for extensive protein engineering.