Stabilization of a Multimeric β-Galactosidase from Thermus sp. Strain T2 by Immobilization on Novel Heterofunctional Epoxy Supports Plus Aldehyde-Dextran Cross-Linking
| Autor: | Cesar Mateo, Alejandro Vian, Alfonso V. Carrascosa, Roberto Fernandez-Lafuente, José Luis García, Benevides C. C. Pessela, Manuel Fuentes, Jose M. Guisan |
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| Jazyk: | angličtina |
| Rok vydání: | 2004 |
| Předmět: |
Immobilized enzyme
Polymers Protein Conformation Lactose Ethylenediamine chemistry.chemical_compound Species Specificity Enzyme Stability Organic chemistry Chelation Thermus Protein Structure Quaternary chemistry.chemical_classification Aldehydes biology Chemistry Hydrolysis Dextrans Enzymes Immobilized beta-Galactosidase biology.organism_classification Combinatorial chemistry Enzyme Activation Dextran Enzyme Covalent bond Epoxy Compounds Protein quaternary structure Adsorption Dimerization Biotechnology |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 1520-6033 8756-7938 |
| Popis: | This 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. This project has been partially funded by CAM (project 07G/0027/2003) |
| Databáze: | OpenAIRE |
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