Precision Engineering of the Co-immobilization of Enzymes for Cascade Biocatalysis.
Angew Chem Int Ed Engl
; 63(22): e202403539, 2024 05 27.
Article
in En
| MEDLINE
| ID: mdl-38556813
ABSTRACT
The design and orderly layered co-immobilization of multiple enzymes on resin particles remain challenging. In this study, the SpyTag/SpyCatcher binding pair was fused to the N-terminus of an alcohol dehydrogenase (ADH) and an aldo-keto reductase (AKR), respectively. A non-canonical amino acid (ncAA), p-azido-L-phenylalanine (p-AzF), as the anchor for covalent bonding enzymes, was genetically inserted into preselected sites in the AKR and ADH. Employing the two bioorthogonal counterparts of SpyTag/SpyCatcher and azide-alkyne cycloaddition for the immobilization of AKR and ADH enabled sequential dual-enzyme coating on porous microspheres. The ordered dual-enzyme reactor was subsequently used to synthesize (S)-1-(2-chlorophenyl)ethanol asymmetrically from the corresponding prochiral ketone, enabling the in situ regeneration of NADPH. The reactor exhibited a high catalytic conversion of 74 % and good reproducibility, retaining 80 % of its initial activity after six cycles. The product had 99.9 % ee, which that was maintained in each cycle. Additionally, the double-layer immobilization method significantly increased the enzyme loading capacity, which was approximately 1.7â
times greater than that of traditional single-layer immobilization. More importantly, it simultaneously enabled both the purification and immobilization of multiple enzymes on carriers, thus providing a convenient approach to facilitate cascade biocatalysis.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Alcohol Dehydrogenase
/
Enzymes, Immobilized
/
Biocatalysis
Language:
En
Journal:
Angew Chem Int Ed Engl
Year:
2024
Document type:
Article
Affiliation country:
Country of publication: