Caracterización cinética de la variante quíntuple mutante de la enzima Petasa de Ideonella sakaiensis

Abstract

Plastic pollution is a global environmental problem, as its production has increased exponentially since its discovery, reaching 470 million tonnes in 2024. Polyethylene terephthalate (PET) is one of the most common plastics, as its thermal and mechanical resistance makes it widely used in the packaging industry. However, these same properties make it difficult to degrade naturally and turn it into a persistent pollutant. Currently, there are several techniques for recovering PET components, although in many cases their quality is lost. Following the discovery of Ideonella sakaiensis and the isolation of PET hydrolases (PETases), biodegradation strategies have focused on developing highly efficient and thermostable PETases for industrial applications. This thesis aims to characterise a quintuple mutant variant capable of efficiently degrading PET. The methodology included esterase activity assays using p-nitrophenyl acetate (pNPA) as an analogue substrate to determine the kinetic constants of the wild-type and mutant variants. Subsequently, their activity was evaluated directly on PET. The results demonstrated that the mutant variant has greater catalytic efficiency compared to the wild-type enzyme, both with the analogue substrate and with PET. In conclusion, this variant represents a significant advance towards the development of sustainable biotechnological processes and their industrial application in the biological recycling of PET.