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RevistaProcess Biochemistry
Año2012
Volumen47
Páginas1220-1227
Internacional

VERSATILITY OF GLUTARALDEHYDE TO IMMOBILIZE LIPASES; EFFECT OF THE IMMOBILIZATION PROTOCOL ON THE PROPERTIES OF LIPASE B FROM Candida antarctica.

Autores:Roberto Fernandez-Lafuente
Grupos de investigación:Optimización de biocatalizadores y bioprocesos enzimáticos
Oveimar Barbosa, Rodrigo Torres,Claudia Ortiz, Roberto Fernandez-Lafuente
 
AFFILIATIONS:  Escuela de Química, Grupo de investigación en Bioquímica y Microbiología (GIBIM), Edificio Camilo Torres 210, Universidad Industrial de Santander, Bucaramanga, Colombia.
Departamento de Biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid. Spain
Escuela de Bacteriología y Laboratorio Clínico, Universidad Industrial de Santander, bucaramanga, Colombia.

ABSTRACT
Glutaraldehyde chemistry has been used to immobilize lipase B from Candida antarctica  (CALB) under different situations. Using high ionic strength, ionic adsorption is avoided, but CALB is adsorbed on the support via interfacial activation. Using non-ionic detergents (e.g., Triton X-100), the enzyme becomes ionically adsorbed on the activated support. If detergent and salt are simultaneously present during immobilization, a covalent attachment to the support is first produced. In absence of detergent or high ionic strength, a mixture of all of the previous immobilization reasons should coexist. Thus, 5 different CALB biocatalysts were prepared following the previous described protocols, and its stability and activity, pH/activity profile and specificity versus R and S methyl mandelate were analyzed. The existence of covalent attachment of more than 95% of the enzyme molecules was confirmed by washing the biocatalysts in salt and detergent solutions. The glutaraldehyde treatment of the enzyme adsorbed on aminated supports did not produce a significant improvement on the activity of the enzyme versus p-nitrophenylpropinate (pNPB) nor a high stabilization of the enzyme. This differed from the effects of a similar treatment of CAL adsorbed on octyl agarose. However, they were similar to the effects of this treatment on covalently immobilized CALB, suggesting that the immobilization protocol may greatly affect the final effect of a chemical modification on the enzyme properties.

Dramatic changes in the enzyme features were observed comparing the different preparations, mainly in the specificity of CALB versus p-NPB and R-methyl mandelate (from 2.5 to 20), or in the enantiospecificity versus R/S methyl mandelate (from 1.8 to 16), confirming that these different immobilization protocols produced biocatalysts with different features. Moreover, changes in experimental conditions produced very different effects on the properties of the different CALB preparations.
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