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PublicationEnzyme and Microbial Technology
Year2018
Volume110
Pages38-45
International

Stabilization of dimeric β-glucosidase from Aspergillus niger via glutaraldehyde immobilization under different conditions

Authors:Roberto Fernandez-Lafuente
Groups of research:Optimization of biocatalysts and bioprocesses
Perla Guadalupe Vazquez-Ortegaa, b, +, Maria Teresa Alcaraz-Fructuosoa,+, Juan A. Rojas-Contrerasb3 , Javier López-Mirandab,*, Roberto Fernandez-Lafuentea,*.
a Departamento de Biocatálisis. ICP-CSIC, Campus UAM-CSIC Madrid, Spain.
b TECNOLÓGICO NACIONAL DE MÉXICO/INSTITUTO TECNOLÓGICO DE DURANGO. Blvd.
8 Felipe Pescador 1130 Ote. Col. Nueva Vizcaya. CP 34080, Durango, Dgo., México.
 
The dimeric enzyme β-glucosidase from Aspergillus niger has been immobilized on different
amino-agarose beads at pH 5 and 7, exploiting the versatility of glutaraldehyde. The stability of the
 free enzyme stability depended on the enzyme concentration. The iImmobilization via ion exchange of
the enzyme improved its enzyme stability/activity, depending on the immobilization pH. However, the
 enzyme was desorbed from the aminated support in 75 mM NaCl at pH 7 and some stability/enzyme
 concentration dependence still existed using this physically immobilized enzyme. Treatment of these
 biocatalysts with glutaraldehyde greatly increased enzyme stability (e.g. at pH 5, after incubation under
conditions where the enzyme just ionically exchanged was fully inactivated, the activity of the
glutaraldehyde treated enzyme remained unaltered). Immobilization on glutaraldehyde pre-activated
supports yielded an evena higher increase in enzyme activity, but the observed stabilization was lower.
While when measuring the enzyme activity at pH 4 there were no clear changes after immobilization,
all immobilized enzymes were clearly more active than the free enzyme at pH 6 and 7 (2-3 times). The
Ki/Km ratio did not significantly enhance decrease in any immobilized biocatalysts, and in some cases
it even worsened in a significant way (by a 9 fold factor using preactivated supports). The new
biocatalysts are significantly more stable and avoid enzyme subunit desorption, andbeing the
immobilization pH is a key point in their design.
 
Keywords:Enzyme immobilization, stabilization of multimeric enzymes, glutaraldehyde versatility, improved enzyme activity upon immobilization, amino-agarose beads, glucosidase
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