Unlocking a Key Residue in a Lipase for Efficient Polyethylene Terephthalate (PET) Hydrolysis and Influencing Depolymerization Product Profiles
Paula Vidal, Ana Robles‐Martín, Laura Fernández-López, José L. González-Alfonso, David Almendral, Rubén Muñoz‐Tafalla, Francisco J. Plou, Vı́ctor Guallar, Manuel Ferrer
Abstract
Abstract Polyethylene terephthalate (PET) pollution is a global challenge. Advancing the bioprospecting of PET‐degrading enzymes through metagenomics and using computational and functional methods to identify key positions influencing the catalytic rate and selectivity are part of the solution. Here, we report PETase activity in the metagenomic lipase Lip MRD9 , which exhibits peak activity at 30 °C and pH 9.0 and has a denaturation temperature of 42 °C. In addition to acting on long‐chain triglycerides (up to ∼13 units (U)/mg, pH 8.0, 30 °C) and a wide range of 34 other esters (up to ∼228 U/g), Lip MRD9 hydrolyses mono(2‐hydroxyethyl) terephthalate (∼57 U/g) and bis(2‐hydroxyethyl) terephthalate (∼131 U/g). It also efficiently deconstructs GoodFellow amorphous submicro‐ and nanosized PET particles (∼984/2238 μM degradation products at 30/40 °C, pH 7.0, 21.5 h) and films (∼112/198 μM degradation products at 30/40 °C, pH 7.0, 7 days). Through molecular modelling and experimental analysis, the active site of Lip MRD9 was revealed, identifying a key residue contributing to its PETase activity compared with those of its closest homologues. This residue plays a crucial role in determining the distinct profiles of degradation products from PET hydrolysis and should be studied in other PETases for its influence on the catalytic process.