(545c) Direct Observation of Conformational Changes of Hydrolases Adsorbed On Fumed Silica Nanoparticles for Catalysis in Organic Solvents

We have recently developed an immobilization strategy for enzymes on fumed silica nanoparticles. Enzymes are adsorbed from aqueous solution on fumed silica followed by lyophilization. The nanobiocatalysts are tested for catalytic activity in organic solvents. Our methodology has proven useful to increase the catalytic activity of Candida antarctica Lipase B (CALB) and subtilisin Carlsberg in hexane by nearly three orders of magnitude compared to the native enzyme suspended in the same solvent. We previously investigated the effect of support surface coverage by the enzyme molecules on the catalytic activity. Here we show some evidence that intricate enzyme-enzyme and enzyme-support interactions produce a maximum in catalytic competency at an intermediate surface coverage. We will show the kinetics of tertiary conformation as inferred from tryptophan fluorescence measurements for a variety of surface coverages, 2,2,2-trifluoroethanol, Dithiothreitol and enzyme concentrations. Circular Dichroism (CD) and Infrared (IR) spectra were collected and deconvoluted to monitor the extent of secondary conformational changes. As external control, the same experiments were conducted for the Lipase of Thermomyces lanuginosus. Relating the physical changes of the enzyme molecules to catalytic competency opens avenues for qualitative prediction of optimum immobilization protocols.