title: Hydrolysis of tannic acid catalyzed by immobilized-stabilized derivatives of Tannase from Lactobacillus plantarum.
abstract: A recombinant tannase from Lactobacillus plantarum , overexpressed in Escherichia coli , was purified in a single step by metal chelate affinity chromatography on poorly activated nickel supports. It was possible to obtain 0.9 g of a pure enzyme by using only 20 mL of chromatographic support. The pure enzyme was immobilized and stabilized by multipoint covalent immobilization on highly activated glyoxyl agarose. Derivatives obtained by multipoint and multisubunit immobilization were 500- and 1000-fold more stable than both the soluble enzyme and the one-point-immobilized enzyme in experiments of thermal and cosolvent inactivation, respectively. In addition, up to 70 mg of pure enzyme was immobilized on 1 g of wet support. The hydrolysis of tannic acid was optimized by using the new immobilized tannase derivative. The optimal reaction conditions were 30% diglyme at pH 5.0 and 4 degrees C. Under these conditions, it was possible to obtain 47.5 mM gallic acid from 5 mM tannic acid as substrate. The product was pure as proved by HPLC. On the other hand, the immobilized biocatalyst preserved >95% of its initial activity after 1 month of incubation under the optimal reaction conditions.
authors.affiliation: Instituto de Fermentaciones Industriales, CSIC, 28006 Madrid, Spain.
authors.forename: Jose Antonio
authors.initials: JA
authors.lastname: Curiel
authors.forename: Lorena
authors.initials: L
authors.lastname: Betancor
authors.forename: Blanca
authors.initials: B
authors.lastname: de las Rivas
authors.forename: Rosario
authors.initials: R
authors.lastname: Muñoz
authors.forename: Jose M
authors.initials: JM
authors.lastname: Guisan
authors.forename: Gloria
authors.initials: G
authors.lastname: Fernández-Lorente
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006868:Hydrolysis; D048191:Lactiplantibacillus plantarum / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D013634:Tannins / Q000378:metabolism*
chemical_list: D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D013634:Tannins; D002265:Carboxylic Ester Hydrolases; C022711:tannase