Journal
ENZYME AND MICROBIAL TECHNOLOGY
Volume 110, Issue -, Pages 38-45Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.enzmictec.2017.12.007
Keywords
Stabilization of multimeric enzymes; Glutaraldehyde versatility; Enzyme immobilization; Improved enzyme activity upon immobilization; Amino-agarose beads; Glucosidase
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Funding
- MINECO from Spanish Government [CTQ2013-41507-R, CTQ2017-86170-R]
- TeNM [DGO-2006-C01-43798]
- National Council of Science and Technology (CONACYT, Mexico) [246459, 291212]
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The dimeric enzyme beta-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 depended on enzyme concentration. Immobilization via ion exchange improved enzyme stability/activity, depending on the immobilization pH. However, the enzyme was desorbed in 75 mM NaCl at pH 7 and some stability/enzyme concentration dependence still existed. Treatment: of these biocatalysts with glutaraldehyde 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 a higher increase in enzyme activity, but the stabilization was lower. While when measuring the enzyme activity at pH 4 there were no changes after immobilization, all immobilized enzymes were more active than the free enzyme at pH 6 and 7 (2-3 times). The Ki/Km ratio did not significantly decrease in any immobilized biocatalysts, and in some cases it 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, being the immobilization pH a key point in their design.
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