title: Glutathione transferases immobilized on nanoporous alumina: flow system kinetics, screening, and stability.
abstract: The previously uncharacterized Drosophila melanogaster Epsilon-class glutathione transferases E6 and E7 were immobilized on nanoporous alumina. The nanoporous anodized alumina membranes were derivatized with 3-aminopropyl-triethoxysilane, and the amino groups were activated with carbonyldiimidazole to allow coupling of the enzymes via ε-amino groups. Kinetic analyses of the immobilized enzymes were carried out in a circulating flow system using CDNB (1-chloro-2,4-dinitrobenzene) as substrate, followed by specificity screening with alternative substrates. A good correlation was observed between the substrate screening data for immobilized enzyme and corresponding data for the enzyme in solution. A limited kinetic study was also carried out on immobilized human GST S1-1 (also known as hematopoietic prostaglandin D synthase). The stability of the immobilized enzymes was virtually identical to that of enzymes in solution, and no leakage of enzyme from the matrix could be observed.
authors.affiliation: Department of Chemistry-BMC, Uppsala University, SE-75123 Uppsala, Sweden.
authors.forename: Marcus
authors.initials: M
authors.lastname: Kjellander
authors.affiliation: Department of Neurochemistry, Stockholm University, SE-10691 Stockholm, Sweden.
authors.forename: Aslam M A
authors.initials: AM
authors.lastname: Mazari
authors.affiliation: Department of Chemistry-Ångström, Uppsala University, SE-75121 Uppsala, Sweden.
authors.forename: Mats
authors.initials: M
authors.lastname: Boman
authors.affiliation: Department of Chemistry-BMC, Uppsala University, SE-75123 Uppsala, Sweden; Department of Neurochemistry, Stockholm University, SE-10691 Stockholm, Sweden.
authors.forename: Bengt
authors.initials: B
authors.lastname: Mannervik
authors.affiliation: Department of Chemistry-BMC, Uppsala University, SE-75123 Uppsala, Sweden. Electronic address: gunnar.johansson@kemi.uu.se.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Johansson
mesh_terms: D000537:Aluminum Oxide / Q000737:chemistry*; D057075:Enzyme Assays / Q000379:methods*; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism*; D005982:Glutathione Transferase / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008567:Membranes, Artificial; D049329:Nanostructures / Q000737:chemistry*; D016062:Porosity
chemical_list: D004800:Enzymes, Immobilized; D008567:Membranes, Artificial; D005982:Glutathione Transferase; D000537:Aluminum Oxide
keywords: Drosophila; Enzyme reactor; Glutathione transferase; Immobilization; Kinetics; Screening