ISBN 9783843941587

978-3-8439-4158-7, Reihe Thermodynamik

Anton Wangler
Prediction and Experimental Validation of Co-Solvent Influences on Enzyme-Catalyzed Reactions

150 Seiten, Dissertation Technische Universität Dortmund (2019), Softcover, A5

Zusammenfassung / Abstract

Enzyme-catalyzed reactions have become significant and viable alternatives to chemically-catalyzed reactions. While enzyme-catalyzed reactions enable catalysis under mild reaction conditions, application is often limited by the low stability of the enzyme. The resulting narrow process window can lead to low reaction yields and slow reaction rates, making the enzyme-catalyzed route unattractive for industrial applications. A way to overcome these limitations is seen in the addition of co-solvents in order to stabilize the enzyme, increase the reaction yield and accelerate the reaction rate. As experimental co-solvent screening is time consuming and expensive, a thermodynamic framework to predict co-solvent influences on reaction equilibrium (and thus the product yield) and reaction kinetics is required. In this work, the co-solvent influence on the reaction equilibrium and the reaction rate of four enzyme-catalyzed reactions was investigated. Two types of reactions were studied, namely a model peptide hydrolysis catalyzed by α-chymotrypsin and the reduction of three different ketones catalyzed by alcohol dehydrogenase. In a first step, the reaction equilibrium and the enzyme-specific kinetic constants, namely the Michaelis constant and the catalytic constant were determined experimentally under neat (co solvent free) reaction conditions and afterwards under the influence of different co solvents. It was found that the addition of co-solvents drastically influences the equilibrium position and the reaction kinetics of all reactions studied. To quantify these influences, experimentally determined reaction equilibrium data and reaction kinetic data under neat conditions were combined with ePC-SAFT predicted activity coefficients of the reacting agents and the enzyme in order to determine thermodynamic equilibrium constants and activity-based kinetic constants. Access to these activity based constants did not just allow quantification of the co-solvent effect, but also enabled predictions of the co-solvent influence, showing excellent agreement with experimental data. Additionally, activity-based kinetic constants were the key to distinguish between enzyme/co-solvent and co-solvent/substrate interactions.