Expedited Award for Novel Research: Enzyme Biocatalysis in Reverse Micelles: Thermodynamic and Kinetic Studies

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The wider use of enzymes in organic syntheses, where their high degree of regio- and stereo-selectivity is difficult to achieve with conventional catalysts, is severely hampered by the low aqueous solubility of many substrates of interest. The encapsulation of enzymes in reverse micelles, which provides a method for stabilization of the enzyme, is a new and particularly promising approach for biological synthesis or transformation of organic compounds in organic solvents. This project concerns fundamental studies for the development of new bioreactors employing such encapsulated enzymes. It includes a study of the effect of encapsulation of dopamine beta- monooxygenase, a non-specific enzyme which hydroxylates ring- substituted phenethylamines, alkenes, aldehydes, amides and sulphides. The effect of micelle structure on the conformation of the enzyme is studied by use of ESR and kinetic measurements. A molecular- thermodynamic model, relating the properties of the aqueous miniphase to prevailing conditions, is also developed. The use of reverse micelles to protect the enzyme from the often denaturing influence of organic solvents is a new approach to enzyme stabilization. Current research has focussed on the use of reverse micelles for protein purification. The inner water pool of a reverse micelle can solubilize large protein molecules and the micelle provides a large surface area for transport of reactant and substrate to and from the enzyme. Little is understood of the behavior of enzymes within such micelles or of how their kinetic properties differ from enzymes in solution. The investigators create micelles using non-polar surfactants, so that transport of polar organic substrates can be enhanced by the addition of carrier molecules in the organic phase, such as quaternary alkyl amines. There is current interest in describing the behavior of water in reverse micelles from a phase equilibrium thermodynamic viewpoint. Of even greater importance is the development of a molecular-thermodynamic model to relate the properties of the aqueous miniphase to the external conditions, so that a predictive model can be employed for bioreactor design.