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RevistaBBA PROTEINS AND PROTEOMICS
Año2019
Volumen1867
Páginas741–747
Internacional

Immobilization of lipase from Pseudomonas fluorescens on glyoxyl-octyl-agarose beads: improved stability and reusability

Autores:Roberto Fernandez-Lafuente
Grupos de investigación:Optimización de biocatalizadores y bioprocesos enzimáticos
Nathalia S. Rios1,2, Carmen Mendez-Sanchez1, Sara Arana-Peña1, Nazzoly Rueda 1,3,  Claudia Ortiz3, Luciana R. B. Gonçalves2 and Roberto Fernandez-Lafuente1,*
 
1Departamento de Biocatálisis. ICP-CSIC, Campus UAM-CSIC, Madrid, Spain.
2Departamento de Engenharia Química, Universidade Federal do Ceará, Campus do Pici, Bloco 709, CEP 60455-760, Fortaleza, CE, Brazil
3Escuela de Microbiología, Universidad Industrial de Santander, Bucaramanga, Colombia.
 
 
Abstract
            The lipase from Pseudomonas fluorescens (PFL) has been immobilized on glyoxyl-octyl agarose and compared to the enzyme immobilized on octyl-agarose. Thus, PFL was immobilized at pH 7 on glyoxyl-octyl support via lipase interfacial activation and later incubated at pH 10.5 for 20 h before reduction to get some enzyme-support covalent bonds. This permitted for 70% of the enzyme molecules to become covalently attached to the support. This biocatalyst was slightly more stable than the octyl-PFL at pH 5, 7 and 9, or in the presence of some organic solvents (stabilization factor no higher than 2). The presence of phosphate anions produced enzyme destabilization, partially prevented by the immobilization on glyoxyl-octyl (stabilization factor became 4). In contrast, the presence of calcium cations promoted a great PFLstabilization, higher in the case of the glyoxyl-octyl preparation (that remained 100% active when the octyl-PFL preparations had lost 20% of the activity). However, it is in the operational stability where the new biocatalyst showed the advantages: in the hydrolysis of 1 M triacetin in 60% 1.4 dioxane, the octyl biocatalyst released more than 60% of the enzyme in the first cycle, while the covalently attached enzyme retained its full activity after 5 reaction cycles.
 
Palabras clave:covalent immobilization, heterofunctional supports, lipase interfacial activation, enzyme release, triacetin hydrolysis
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