Modificación química dirigida de lipasas en fase sólida

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Química Analítica y Ánalisis Instrumental. Fecha de lectura: 25-02-2014 ; Enzymes are versatile biocatalysts finding an increasing application in many industrial fields including fine and organic chemistry, pharmacy, cosmetics or food industry. The enzymes unsurpassed chemo-, regioand enantioselectivity characteristics are making them interesting as fine chemistry biocatalysts since the preparation of pure intermediates is a key step for many industrially relevant processes. However, when using non-natural substrates enzyme specificity may not be as high as required for industrial purposes requesting the need of improvement of this enzyme property, possible through very different techniques, like screening, controlled immobilization, enzyme engineering, directed evolution and rational design approaches, chemical modification or combinations thereof. Strategies for the selective and efficient chemical modification of proteins have been implemented successfully for the study and modulation of enzyme specificity. However, despite the efforts made the past years the site-directed incorporation of different moieties, like carbohydrates, peptides, polymers, DNA, etc. specifically to the required position at the enzyme remains a challenging task. Applying conventional methods of chemical modification on a protein attached covalently or reversible to a surface or matrix can provide an important simplification of current protocols of modification. The main objective of this Doctoral Thesis has been the development and implementation of new strategies of site-directed chemical modifications of proteins on solid phase to improve their catalytic properties. Joining genetic and chemical modification approaches 5 strategies were developed using immobilized lipases as model enzymes: - Chemical glycosylation of lipases on solid phase - Promotion of the open conformation o lipase by two cysteine ligation - Development of ...