title: Reversible covalent immobilization of Trametes villosa laccase onto thiolsulfinate-agarose: An insoluble biocatalyst with potential for decoloring recalcitrant dyes.
abstract: The development of a solid-phase biocatalyst based on the reversible covalent immobilization of laccase onto thiol-reactive supports (thiolsulfinate-agarose [TSI-agarose]) was performed. To achieve this goal, laccase-producing strains isolated from Eucalyptus globulus were screened and white rot fungus Trametes villosa was selected as the best strain for enzyme production. Reduction of disulfide bonds and introduction of "de novo" thiol groups in partially purified laccase were assessed to perform its reversible covalent immobilization onto thiol-reactive supports (TSI-agarose). Only the thiolation process dramatically improved the immobilization yield, from 0% for the native and reduced enzyme to 60% for the thiolated enzyme. Mild conditions for the immobilization process (pH 7.5 and 4°C) allowed the achievement of nearly 100% of coupling efficiency when low loads were applied. The kinetic parameters, pH, and thermal stabilities for the immobilized biocatalyst were similar to those for the native enzyme. After the first use and three consecutives reuses, the insoluble derivative kept more than 80% of its initial capacity for decolorizing Remazol Brilliant Blue R, showing its suitability for color removal from textile industrial effluents. The possibility of reusing the support was demonstrated by the reversibility of enzyme-support binding.
authors.affiliation: Cátedra de Bioquímica, Departamento de Biociencias, Facultad de Química, Universidad de la República, Montevideo, Uruguay.
authors.forename: Larissa
authors.initials: L
authors.lastname: Gioia
authors.affiliation: CEIT, Unit of Environmental Engineering, San Sebastian 20018, Spain.|IKERBASQUE, Basque Foundation for Science, Bilbao 48011, Spain.
authors.forename: Susana
authors.initials: S
authors.lastname: Rodríguez-Couto
authors.affiliation: Laboratorio de Biocatálisis y Biotransformaciones, Departamento de Química Orgánica, Facultad de Química, Universidad de la República, Montevideo, Uruguay.
authors.forename: María Del Pilar
authors.initials: Mdel P
authors.lastname: Menéndez
authors.affiliation: Cátedra de Bioquímica, Departamento de Biociencias, Facultad de Química, Universidad de la República, Montevideo, Uruguay.
authors.forename: Carmen
authors.initials: C
authors.lastname: Manta
authors.affiliation: Cátedra de Bioquímica, Departamento de Biociencias, Facultad de Química, Universidad de la República, Montevideo, Uruguay.
authors.forename: Karen
authors.initials: K
authors.lastname: Ovsejevi
mesh_terms: D000880:Anthraquinones / Q000737:chemistry*; D002384:Catalysis; D004396:Coloring Agents / Q000737:chemistry*; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005656:Fungal Proteins / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D042845:Laccase / Q000737:chemistry*; D012685:Sepharose / Q000737:chemistry*; D055454:Trametes / Q000201:enzymology*
chemical_list: D000880:Anthraquinones; D004396:Coloring Agents; D004800:Enzymes, Immobilized; D005656:Fungal Proteins; D012685:Sepharose; D042845:Laccase; C024758:Remazol Brilliant Blue R
keywords: Remazol Brilliant Blue R; Trametes villosa; covalent reversible immobilization; laccase; thiolsulfinate-agarose; white rot fungi