Three in One : Temperature, Solvent and Catalytic Stability by Engineering the Cofactor-Binding Element of Amine Transaminase
Börner, Tim LU ; Rämisch, Sebastian LU ; Bartsch, Sebastian ; Vogel, Andreas ; Adlercreutz, Patrick LU
and Grey, Carl
LU
(2017)
In ChemBioChem
18(15).
p.1482-1486
- Abstract
Amine transaminase (ATA) catalyse enantioselectively the direct amination of ketones, but insufficient stability during catalysis limits their industrial applicability. Recently, we revealed that ATAs suffer from substrate-induced inactivation mechanism involving dissociation of the enzyme-cofactor intermediate. Here, we report on engineering the cofactor-ring-binding element, which also shapes the active-site entrance. Only two point mutations in this motif improved temperature and catalytic stability in both biphasic media and organic solvent. Thermodynamic analysis revealed a higher melting point for the enzyme-cofactor intermediate. The high cofactor affinity eliminates the need for pyridoxal 5′-phosphate supply, thus making... (More)
Amine transaminase (ATA) catalyse enantioselectively the direct amination of ketones, but insufficient stability during catalysis limits their industrial applicability. Recently, we revealed that ATAs suffer from substrate-induced inactivation mechanism involving dissociation of the enzyme-cofactor intermediate. Here, we report on engineering the cofactor-ring-binding element, which also shapes the active-site entrance. Only two point mutations in this motif improved temperature and catalytic stability in both biphasic media and organic solvent. Thermodynamic analysis revealed a higher melting point for the enzyme-cofactor intermediate. The high cofactor affinity eliminates the need for pyridoxal 5′-phosphate supply, thus making large-scale reactions more cost effective. This is the first report on stabilising a tetrameric ATA by mutating a single structural element. As this structural "hotspot" is a common feature of other transaminases it could serve as a general engineering target.
(Less)
- author
- Börner, Tim
LU
; Rämisch, Sebastian
LU
; Bartsch, Sebastian
; Vogel, Andreas
; Adlercreutz, Patrick
LU
and Grey, Carl
LU
- organization
- publishing date
- 2017-08-04
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Amines, Enzyme catalysis, Operational stability, Pyridoxamine 5′-phosphate, Transaminase
- in
- ChemBioChem
- volume
- 18
- issue
- 15
- pages
- 1482 - 1486
- publisher
- John Wiley & Sons Inc.
- external identifiers
-
- scopus:85020430872
- pmid:28470825
- wos:000406932800004
- ISSN
- 1439-4227
- DOI
- 10.1002/cbic.201700236
- language
- English
- LU publication?
- yes
- id
- d99f3583-b612-43fc-8ed6-e23670d09fdc
- date added to LUP
- 2017-06-30 11:11:42
- date last changed
- 2024-12-17 02:47:23
@article{d99f3583-b612-43fc-8ed6-e23670d09fdc,
abstract = {{<p>Amine transaminase (ATA) catalyse enantioselectively the direct amination of ketones, but insufficient stability during catalysis limits their industrial applicability. Recently, we revealed that ATAs suffer from substrate-induced inactivation mechanism involving dissociation of the enzyme-cofactor intermediate. Here, we report on engineering the cofactor-ring-binding element, which also shapes the active-site entrance. Only two point mutations in this motif improved temperature and catalytic stability in both biphasic media and organic solvent. Thermodynamic analysis revealed a higher melting point for the enzyme-cofactor intermediate. The high cofactor affinity eliminates the need for pyridoxal 5′-phosphate supply, thus making large-scale reactions more cost effective. This is the first report on stabilising a tetrameric ATA by mutating a single structural element. As this structural "hotspot" is a common feature of other transaminases it could serve as a general engineering target.</p>}},
author = {{Börner, Tim and Rämisch, Sebastian and Bartsch, Sebastian and Vogel, Andreas and Adlercreutz, Patrick and Grey, Carl}},
issn = {{1439-4227}},
keywords = {{Amines; Enzyme catalysis; Operational stability; Pyridoxamine 5′-phosphate; Transaminase}},
language = {{eng}},
month = {{08}},
number = {{15}},
pages = {{1482--1486}},
publisher = {{John Wiley & Sons Inc.}},
series = {{ChemBioChem}},
title = {{Three in One : Temperature, Solvent and Catalytic Stability by Engineering the Cofactor-Binding Element of Amine Transaminase}},
url = {{http://dx.doi.org/10.1002/cbic.201700236}},
doi = {{10.1002/cbic.201700236}},
volume = {{18}},
year = {{2017}},
}