Immobilization and stabilization of a cyclodextrin glycosyltransferase by covalent attachment on highly activated glyoxyl-agarose supports.
Covalent immobilization of cyclodextrin glycosyltransferase on glyoxyl-agarose beads promotes a very high stabilization of the enzyme against any distorting agent (temperature, pH, organic solvents). For example, the optimized immobilized preparation preserves 90% of initial activity when incubated for 22 h in 30% ethanol at pH 7 and 40 degrees C. Other immobilized preparations (obtained via other immobilization protocols) exhibit less than 10% of activity after incubation under similar conditions. Optimized glyoxyl-agarose immobilized preparation expressed a high percentage of catalytic activity (70%). Immobilization using any technique prevents enzyme inactivation by air bubbles during strong stirring of the enzyme. Stabilization of the enzyme immobilized on glyoxyl-agarose is higher when using the highest activation degree (75 micromol of glyoxyl per milliliter of support) as well as when performing long enzyme-support incubation times (4 h) at room temperature. Multipoint covalent immobilization seems to be responsible for this very high stabilization associated to the immobilization process on highly activated glyoxyl-agarose. The stabilization of the enzyme against the inactivation by ethanol seems to be interesting to improve cyclodextrin production: ethanol strongly inhibits the enzymatic degradation of cyclodextrin while hardly affecting the cyclodextrin production rate of the immobilized-stabilized preparation.
Ferrarotti
Susana Alicia
SA
Departamento de Biocatalisis, Instituto de Catalisis-CSIC, Campus UAM, Cantoblanco, Madrid, Spain.
Bolivar
Juan M
JM
Mateo
Cesar
C
Wilson
Lorena
L
Guisan
Jose M
JM
Fernandez-Lafuente
Roberto
R
D004800:Enzymes, Immobilized / Q000737:chemistry*; D000431:Ethanol / Q000737:chemistry; D005964:Glucosyltransferases / Q000737:chemistry*; D006038:Glyoxylates / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D012685:Sepharose / Q000737:chemistry*; D013696:Temperature; D013997:Time Factors
D004800:Enzymes, Immobilized; D006038:Glyoxylates; C070399:glyoxyl agarose; D000431:Ethanol; D012685:Sepharose; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase