title: Use of immobilized lipases for lipase purification via specific lipase-lipase interactions.
abstract: Lipase from Pseudomonas fluorescens (PFL), an enzyme with a great tendency to yield bimolecular aggregates, was immobilized via multipoint covalent attachment on glyoxyl-agarose in the presence of Triton X-100. This strategy permitted to obtain the enzyme with the active center oriented towards the reaction medium. This immobilized enzyme presents the capacity of specifically adsorbing PFL molecules, that can be easily desorbed by the use of detergents. More interesting, the enzyme was also able to adsorb other lipases. That is, the lipase from Bacillus thermocatenulatus (BTL2) cloned in Escherichia coli was selectively adsorbed on this immobilized enzyme, enabling a very simple purification strategy. Similar results were achieved with some other lipases (those from Rhizomucor miehei (RML), Rhizopus oryzae (ROL), and Humicola Lanuginosa (HLL)). In all cases, the enzyme could be easily desorbed by incubation with Triton X-100. The matrix could be used several cycles without any detrimental effect on the adsorption capacity.
authors.affiliation: Departamento of Biocatalisis, Instituto of Catalisis (CSIC), Campus UAM Cantoblanco, Madrid 28049, Spain.
authors.forename: Jose M
authors.initials: JM
authors.lastname: Palomo
authors.forename: Claudia
authors.initials: C
authors.lastname: Ortiz
authors.forename: Manuel
authors.initials: M
authors.lastname: Fuentes
authors.forename: Gloria
authors.initials: G
authors.lastname: Fernandez-Lorente
authors.forename: Jose M
authors.initials: JM
authors.lastname: Guisan
authors.forename: Roberto
authors.initials: R
authors.lastname: Fernandez-Lafuente
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel; D004800:Enzymes, Immobilized / Q000378:metabolism*; D008049:Lipase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D008049:Lipase