title: Effect of protein load on stability of immobilized enzymes.
abstract: Different lipases have been immobilized on octyl agarose beads at 1mg/g and at maximum loading, via physical interfacial activation versus the octyl layer on the support. The stability of the preparations was analyzed. Most biocatalysts had the expected result: the apparent stability increased using the highly loaded preparations, due to the diffusional limitations that reduced the initial observed activity. However, lipase B from Candida antarctica (CALB) was significantly more stable using the lowly loaded preparation than the maximum loaded one. This negative effect of the enzyme crowding on enzyme stability was found in inactivations at pH 5, 7 or 9, but not in inactivations in the presence of organic solvents. The immobilization using ethanol to reduce the immobilization rate had no effect on the stability of the lowly loaded preparation, while the highly loaded enzyme biocatalysts increased their stabilities, becoming very similar to that of the lowly loaded preparation. Results suggested that CALB molecules immobilized on octyl agarose may be closely packed together due to the high immobilization rate and this produced some negative interactions between immobilized enzyme molecules during enzyme thermal inactivation. Slowing-down the immobilization rate may be a solution for this unexpected problem.
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Fernandez-Lopez
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain.
authors.forename: Sara G
authors.initials: SG
authors.lastname: Pedrero
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain.
authors.forename: Nerea
authors.initials: N
authors.lastname: Lopez-Carrobles
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain.
authors.forename: Beatriz C
authors.initials: BC
authors.lastname: Gorines
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain; CONACYT-Centro de Investigación en Alimentación y Desarrollo, A.C., Centro de Innovación y Desarrollo Agroalimentario de Michoacán, A.C. (CIDAM), Km. 8 Antigua Carretera a Pátzcuaro s/n, C.P. 58341 Morelia, Michoacán, Mexico. Electronic address: juanvirgen@hotmail.com.
authors.forename: Jose J
authors.initials: JJ
authors.lastname: Virgen-Ortíz
authors.affiliation: Departamento de biocatálisis. Instituto de Catálisis-CSIC, Campus UAM-CSIC Madrid, Spain. Electronic address: rfl@icp.csic.es.
authors.forename: Roberto
authors.initials: R
authors.lastname: Fernandez-Lafuente
mesh_terms: D001203:Ascomycota / Q000201:enzymology; D055162:Biocatalysis; D001709:Biotechnology; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D005656:Fungal Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008049:Lipase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D020103:Rhizomucor / Q000201:enzymology; D012685:Sepharose / Q000031:analogs & derivatives; D012997:Solvents
chemical_list: D004800:Enzymes, Immobilized; D005656:Fungal Proteins; D012997:Solvents; C027176:octyl-sepharose CL-4B; D012685:Sepharose; D008049:Lipase; C431349:lipase B, Candida antarctica
keywords: Control of immobilization rate; Enzyme crowding; Enzyme stability; Immobilized enzyme interactions; Octyl agarose