Development of organic solvent stable enzymes by protein engineering

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Enzymes are very good catalysts, because they effectively and specifically catalyze many kinds of reactions under mild conditions. Then they do not produce by-products and not consume a lot of energy. When enzyme are used as a catalyst in a chemical process, it is possible to reduce process steps, to increase yield, to reduce raw materials, waste materials, solvents, energy, equipment, utilities, production costs, environment pollution, and to increase profits and competitiveness. However, sometimes some advantages of enzymes become disadvantages. Although enzymes are active under mild condition, they are easily denatured and lose catalytic activity under extreme conditions, such as low pH, high pH and temperature, and also in the presence of organic solvents. Especially, although most fine chemicals are insoluble in water, they are soluble in organic solvents. Then organic solvent-tolerant enzymes are required as catalysts to produce fine chemicals. In this research, clarification of organic solvent-tolerance of enzymes and development of new organic solvent-tolerant enzymes using protein engineering were performed. The examined item in this research is as follows.(1) Understanding of the organic solvent-tolerance of enzyme(2) Gene cloning of the enzyme in which organic solvent tolerance is desired(3) Construction of the gene expression system of the organic solvent tolerant enzyme(4) Development of the organic solvent tolerant enzyme having useful substrate specificity by the protein engineering