PRODUCTION METHOD OF NITRILASE AND NITRILEHYDRATASE WHICH CONTAINS IMMOBILIZATION THEREOF

摘要

PURPOSE: A method for producing nitrilase and nitrilehydratase is provided to ensure high enzyme purity and to reduce production cost. CONSTITUTION: A method for producing enzyme comprises: a step of adding surfactant to recombinant E.coli and isolating enzyme; a step of adding 5-30 wt% of 3-aminopropyl triethoxy silane and silanizing a carrier; a step of treating the carrier with glutaraldehyde; and a step of adding an enzyme extract to the carrier and fixing the enzyme. The enzyme is nitrilase or nitrilehyratase. The carrier is a silica gel carrier. [Reference numerals] (AA) Carrier; (BB) Silanization (adding 3-aminopropyl-triethoxy silane and stirring at 90°C for 2-3 hours; (CC) Drying (105°C, 2-3 hours); (DD) Washing with distilled water(90°C, 1 hour); (EE) Adding 50% glutaraldehyde to dried carrier; (FF) Stirring(room temperature, 1-2 hours); (GG, LL, OO) Washing with distilled water; (HH) Adding 200u/ml of each concentrated nitrilase and Nitrile hydratase and suspending with 0.1M potassium phosphate buffer(pH 7.5); (II) Stirring(room temperature, 2-4 hours); (JJ) Adding L-lysine(0.5% of dried carrier weight); (KK,NN) Stirring(room temperature, 1 hour); (MM) Adding 1N NaCl(5 times volume of the dried carrier); (PP) Washing with 1M Potassium phosphate buffer(pH7.5)