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Development of a More Stable and Methanol Tolerant Lipase for Improved Production of Biodiesel Products

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Summary: Biodiesel is an attractive alternative to petrodiesel since it burns cleaner releases less CO2 is biodegradable and can be obtained from renewable sources. Additionally since biodiesel is compatible with current diesel engines there is a large potential market for biodiesel to enter. Current methods for biodiesel synthesis include base-catalyzed reactions and lipase-catalyzed reactions. Both of these reactions produce biodiesels in the form of fatty acid methyl esters (FAMEs) as well as glycerol which can also be purified and sold as a value-added product. While bases are an inexpensive catalyst base-catalyzed reactions lead to increased production of unwanted contaminates that require additional purification to yield a final product. In contrast lipase-catalyzed reactions do not produce such contaminants and are more easily isolated from the final products. However natural lipases are unstable at high temperatures and irreversibly inactivated at high methanol concentrations used in biodiesel production limiting their practical use. Therefore in order to utilize the more favorable lipase-catalyzed reactions the development of a lipase enzyme with high enzymatic activity and stability at high temperatures in the presence of methanol is needed. Detailed Technology Description: Dr. James Bowie Dr. Tyler Korman and colleagues in UCLA’s Department of Chemistry and Biochemistry have engineered a highly stable and methanol tolerant lipase through directed evolution. The engineered lipase (Dieselzyme 4) is a variant of the natural lipase found in P. mirabilus. Dieselzyme 4 has 30-fold increased stability over the wild type enzyme at 50°C and 50-fold increased stability at high methanol concentrations. Additionally Dieselzyme 4 retains its ability synthesize biodiesel through many cycles of synthesis demonstrating improved longevity over both the wild type enzyme and an industry standard lipase (B. cepacia). The improved methanol tolerance thermostability and longevity of Dieselzyme 4 make it easily the most robust lipase currently available for biodiesel production. Unlike many other lipases Dieselzyme 4 is easily expressed in E. coli allowing for ease of large-scale production. The ease of production also makes it an ideal platform for further engineering efforts in the future. The inventors are particularly interested in collaborating with industrial partners to develop further enhanced Dieselzymes. Technology Applications: •Replacement of natural lipases currently in use in biodiesel production •Platform for the development of a further improved enzyme State Of Development: Dieselzyme 4 was engineered through directed evolution. The enzyme has demonstrated increased stability and enzymatic activity at high temperatures and methanol concentrations. It has also demonstrated the ability to catalyze many rounds of biodiesel production.


•Increased enzymatic activity in the presence of methanol •Increased thermostability •Can catalyze many rounds of biodiesel synthesis •Can be easily produced in high quantities using E. coli

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