Enhanced transglucosylation/hydrolysis ratio of mutants of Pyrococcus furiosus β-glucosidase : Effects of donor concentration, water content, and temperature on activity and selectivity in hexanol
(2001) In Biotechnology and Bioengineering 75(6). p.656-665- Abstract
The transglucosylation reaction catalyzed by wild-type β-glucosidase CelB from hyperthermophilic Pyrococcus furiosus and active site mutants (M424K, F426Y, M424K/F426Y) was studied. The conversion of pentyl-β-glucoside to hexyl-β-glucoside in hexanol was used as a model transglucosylation reaction. Hydrolysis to glucose was a side reaction. The activity (rates of hydrolysis and transglucosylation) and the selectivity (S value) were measured as a function of pentyl-β-glucoside concentration (5-240 mM), water content (1-100% v/v), and temperature (50-95°C). All mutants had lower activity than the wild-type enzyme, but they had higher selectivity, which means that they provided a higher ratio of transglucosylation product to hydrolysis... (More)
The transglucosylation reaction catalyzed by wild-type β-glucosidase CelB from hyperthermophilic Pyrococcus furiosus and active site mutants (M424K, F426Y, M424K/F426Y) was studied. The conversion of pentyl-β-glucoside to hexyl-β-glucoside in hexanol was used as a model transglucosylation reaction. Hydrolysis to glucose was a side reaction. The activity (rates of hydrolysis and transglucosylation) and the selectivity (S value) were measured as a function of pentyl-β-glucoside concentration (5-240 mM), water content (1-100% v/v), and temperature (50-95°C). All mutants had lower activity than the wild-type enzyme, but they had higher selectivity, which means that they provided a higher ratio of transglucosylation product to hydrolysis product. The largest increase in S-value (2.6 fold) was obtained by the F426Y mutant, which resulted in increased hexyl-β-glucoside yield from 56% to 69%. In addition, the F426Y enzyme had higher selectivity over the wide range of temperatures tested. The activity of CelB wild-type and CelB F426Y increased as a function of water activity (aw), and complete activation by the water was obtained in a two-phase system with 20% water phase. In contrast to CelB wild-type, the F426Y mutant had transferase activity as low as aw = 0.29. Surprisingly, the S value increased with increasing water activity up to aw = 0.92. At still higher water content the S value decreased.
(Less)
- author
- Hansson, Therese
LU
and Adlercreutz, Patrick
LU
- organization
- publishing date
- 2001-12-20
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- βglucosidase, Activity, Hexanol, Selectivity, Transglucosylation
- in
- Biotechnology and Bioengineering
- volume
- 75
- issue
- 6
- pages
- 10 pages
- publisher
- John Wiley & Sons Inc.
- external identifiers
-
- pmid:11745143
- scopus:0035924115
- ISSN
- 0006-3592
- DOI
- 10.1002/bit.10043
- language
- English
- LU publication?
- yes
- id
- 5ab4523a-b7dd-45a5-b73a-7e1c79fea249
- date added to LUP
- 2019-06-20 15:48:18
- date last changed
- 2025-04-04 14:22:30
@article{5ab4523a-b7dd-45a5-b73a-7e1c79fea249, abstract = {{<p>The transglucosylation reaction catalyzed by wild-type β-glucosidase CelB from hyperthermophilic Pyrococcus furiosus and active site mutants (M424K, F426Y, M424K/F426Y) was studied. The conversion of pentyl-β-glucoside to hexyl-β-glucoside in hexanol was used as a model transglucosylation reaction. Hydrolysis to glucose was a side reaction. The activity (rates of hydrolysis and transglucosylation) and the selectivity (S value) were measured as a function of pentyl-β-glucoside concentration (5-240 mM), water content (1-100% v/v), and temperature (50-95°C). All mutants had lower activity than the wild-type enzyme, but they had higher selectivity, which means that they provided a higher ratio of transglucosylation product to hydrolysis product. The largest increase in S-value (2.6 fold) was obtained by the F426Y mutant, which resulted in increased hexyl-β-glucoside yield from 56% to 69%. In addition, the F426Y enzyme had higher selectivity over the wide range of temperatures tested. The activity of CelB wild-type and CelB F426Y increased as a function of water activity (a<sub>w</sub>), and complete activation by the water was obtained in a two-phase system with 20% water phase. In contrast to CelB wild-type, the F426Y mutant had transferase activity as low as a<sub>w</sub> = 0.29. Surprisingly, the S value increased with increasing water activity up to a<sub>w</sub> = 0.92. At still higher water content the S value decreased.</p>}}, author = {{Hansson, Therese and Adlercreutz, Patrick}}, issn = {{0006-3592}}, keywords = {{βglucosidase; Activity; Hexanol; Selectivity; Transglucosylation}}, language = {{eng}}, month = {{12}}, number = {{6}}, pages = {{656--665}}, publisher = {{John Wiley & Sons Inc.}}, series = {{Biotechnology and Bioengineering}}, title = {{Enhanced transglucosylation/hydrolysis ratio of mutants of Pyrococcus furiosus β-glucosidase : Effects of donor concentration, water content, and temperature on activity and selectivity in hexanol}}, url = {{http://dx.doi.org/10.1002/bit.10043}}, doi = {{10.1002/bit.10043}}, volume = {{75}}, year = {{2001}}, }