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Title

Stabilization of a Multimeric β-Galactosidase from Thermus sp. Strain T2 by Immobilization on Novel Heterofunctional Epoxy Supports Plus Aldehyde-Dextran Cross-Linking

AuthorsPessela, Benevides C. ; Mateo, César; Fuentes, Manuel ; Vian Herrero, Alejandro; García, José Luis ; Carrascosa, Alfonso V. ; Guisán, José M.; Fernández-Lafuente, Roberto
Issue Date2004
PublisherAmerican Institute of Chemical Engineers
CitationBiotechnology Progress 20(1):388-392(2004)
AbstractThis work exemplifies the advantages of using a battery of new heterofunctional epoxy supports to immobilize enzymes. We have compared the performance of a standard Sepabeads-epoxy support with other Sepabeads-epoxy supports partially modified with boronate, iminodiacetic, metal chelates, and ethylenediamine in the immobilization of the thermostable β-galactosidase from Thermus sp. strain T2 as a model system. Immobilization yields depended on the support, ranging from 95% using Sepabeads-epoxy-chelate, Sepabeads-epoxy-amino, or Sepabeads-epoxy-boronic to 5% using Sepabeads-epoxy-IDA. Moreover, immobilization rates were also very different when using different supports. Remarkably, the immobilized β-galactosidase derivatives showed very improved but different stabilities after favoring multipoint covalent attachment by long-term alkaline incubation, the enzyme immobilized on Sepabeads-epoxy-boronic being the most stable. This derivative had some subunits of the enzyme not covalently attached to the support (detected by SDS-PAGE). This is a problem if the biocatalysts were to be used in food technology. The optimization of the cross-linking with aldehyde-dextran permitted the full stabilization of the quaternary structure of the enzyme. The optimal derivative was very active in lactose hydrolysis even at 70°C (over 1000 IU/g), maintaining its activity after long incubation times under these conditions and with no risk of product contamination with enzyme subunits.
URIhttp://hdl.handle.net/10261/73796
DOI10.1021/bp034183f
Identifiersdoi: 10.1021/bp034183f
issn: 8756-7938
e-issn: 1520-6033
Appears in Collections:(CIB) Artículos
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