Immobilization of Alcohol Dehydrogenase from E. coli onto Mesoporous Silica for Application as a Cofactor Recycling System

AbstractAlcohol dehydrogenase (ADH) from Escherichia coli (E. coli) was electrostatically immobilized onto the surface of aminopropyl modified mesoporous cellular siliceous foams (MCF‐NH2). To ascertain the optimal pH for the immobilization, several pH values (pH 6.5, 7.0, 7.5) were examined. The MCF‐NH2 amount applied for immobilization was also varied to identify the most suitable enzyme to host ratio. ADH uptakes, long‐term stabilities, and leaching behavior were monitored, and the forward and back reactions were studied. ADH activity after 24 h of immobilization decreased with increasing pH. Independent of the pH, the best long‐term stabilities were obtained with an enzyme to silica ratio of 1:10. The back reaction is always preferred and is thus much faster. Michaelis–Menten kinetics of the free and the immobilized ADH of the forward as well as the back reaction were elaborated for each immobilization pH and by employing an ADH to MCF‐NH2 ratio of 1:10. Turnover numbers and the enzyme efficiencies (EE) were determined. From the kinetic aspects the immobilization was most promising when using a pH 6.5 buffer (forward reaction) or pH 7.0 buffer (back reaction) for immobilization. Thus, depending on the coenzyme that needs to be recycled (NADP+ or NADPH) the pH of the immobilization has to be selected.