Modulation of a lipase from Staphylococcus warneri EX17 using immobilization techniques |
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Authors: | Giandra Volpato, Marco Filice, Rafael C. Rodrigues, Jú lio X. Heck, Jose M. Guisan, Cesar Mateo,Marco Ant nio Z chia Ayub |
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Affiliation: | aDepartamento de Biocatálisis, Instituto de Catálisis-CSIC, Campus UAM, Cantoblanco, 28049 Madrid, Spain;bFood Science and Technology Institute, Federal University of Rio Grande do Sul State, Av. Bento Gonçalves, 9500, P.O. Box 15090, ZC 91501-970, Porto Alegre, RS, Brazil;cTechnical School, Federal University of Rio Grande do Sul State, Rua Ramiro Barcelos, 2777, ZC 90035-007 Porto Alegre, RS, Brazil |
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Abstract: | This research describes the immobilization on glyoxyl, cyanogen bromide or octyl agarose beads of a purified lipase from Staphylococcus warneri strain EX17 (SWL), and the effect on its properties. The immobilization on glyoxyl-agarose at pH 10 and 25 °C, conditions in which the enzyme is readily inactivated, required the stabilization of the soluble enzyme. This was attained by the addition of 25% glycerol. Using this additive, immobilization on glyoxyl-agarose beads proceeded very quickly with good activity retention around 80%. This was the most stable preparation under thermal inactivation at pH 5, 7 and 9, in the presence of either cosolvents or detergents. This preparation was hyperactivated by concentrations of Triton X-100, which would produce negative effects over enzyme activity when using the other SWL preparations. Immobilized SWL preparations hydrolyzed different chiral esters, such as (±)-methyl mandelate, (±)-2-O-butyryl-2-phenylacetic acid, and (±)-2-hydroxy-4-phenyl-butyric acid ethyl ester, being its specificity depended on the immobilization protocol. The enantiospecificity was also strongly modulated by the immobilization. Thus, using HPBEt as substrate, octyl-SWL exhibited an opposite enantiospecificity to the other two biocatalysts. This preparation was the most enantioselective in the hydrolysis of (±)-2-O-butyryl-2-phenylacetic acid (E = 56.3). |
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Keywords: | Modulation of lipases Enzyme stabilization Interfacial activation Hyperactivation by detergents Glyoxyl-agarose |
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