Computational Design and Redesign of Hydrolases for Biotechnological Applications
[eng] Hydrolases, a versatile class of enzymes, play a pivotal role in numerous biotechnological applications, including waste recycling and improved laundry efficiency in the detergent industry. Recent advances in computational approaches have revolutionized the design and optimization of these enz...
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2026 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:dnet:ubarcelona__::edc3f2542ddfbff0ccf1ed9fb3100fca |
| Acceso en línea: | https://hdl.handle.net/2445/229390 https://hdl.handle.net/10803/697446 |
| Access Level: | acceso embargado |
| Palabra clave: | Hidrolases Enzimologia Hydrolases Enzymology |
| Sumario: | [eng] Hydrolases, a versatile class of enzymes, play a pivotal role in numerous biotechnological applications, including waste recycling and improved laundry efficiency in the detergent industry. Recent advances in computational approaches have revolutionized the design and optimization of these enzymes, enabling precise tuning of their activity, and substrate specificity. This thesis highlights the impact of molecular modeling and structure-guided strategies in engineering hydrolases, focusing on both the fine-tuning of natural scaffolds and the creation of de novo catalytic architectures. A metagenomic lipase, LipMRD9, was identified as a dual lipase/PETase and shown to rely on a key residue for productive PET hydrolysis. Guided by physics-based simulations and energy landscape exploration, LipMRD9 was redesigned to yield variants with up to threefold higher PET activity while retaining lipase function. To transcend the limitations of soluble enzymes, pore-forming proteins were engineered into “porezymes” by embedding artificial Ser–His–Asp/Glu catalytic triads within their lumens. These catalytic nanopores displayed hydrolytic activity against PET nanoparticles surpassing benchmark hydrolases under mesophilic conditions, establishing them as a novel class of catalytic pores. Automated computational pipelines extended this principle to large pore scaffolds such as pneumolysin and -hemolysin, demonstrating scalability and potential for multisite catalysis. By integrating structural biology, pluriZyme engineering methodologies, and advanced computational design, this work provides complementary solutions for PET degradation and sets the stage for sustainable, cost-effective applications in industrial biotechnology. Future directions emphasize the convergence of computational design with experimental validation to fully unlock the potential of hydrolases in addressing global environmental and industrial challenges. |
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