Mattias Basle
Design of artificial metalloenzymes for the reduction of nicotinamide cofactors
Basle, Mattias; Padley, Henry A.W.; Martins, Floriane L.; Winkler, Gerlof Sebastiaan; J�ger, Christof M.; Pordea, Anca
Authors
Henry A.W. Padley
Floriane L. Martins
SEBASTIAAN WINKLER SEBASTIAAN.WINKLER@NOTTINGHAM.AC.UK
Associate Professor
Christof M. J�ger
ANCA PORDEA ANCA.PORDEA@NOTTINGHAM.AC.UK
Associate Professor
Abstract
Artificial metalloenzymes result from the insertion of a catalytically active metal complex into a biological scaffold, generally a protein devoid of other catalytic functionalities. As such, their design requires efforts to engineer substrate binding, in addition to accommodating the artificial catalyst. Here we constructed and characterised artificial metalloenzymes using alcohol dehydrogenase as starting point, an enzyme which has both a cofactor and a substrate binding pocket. A docking approach was used to determine suitable positions for catalyst anchoring to single cysteine mutants, leading to an artificial metalloenzyme capable to reduce both natural cofactors and the hydrophobic 1-benzylnicotinamide mimic. Kinetic studies revealed that the new construct displayed a Michaelis-Menten behaviour with the native nicotinamide cofactors, which were suggested by docking to bind at a surface exposed site, different compared to their native binding position. On the other hand, the kinetic and docking data suggested that a typical enzyme behaviour was not observed with the hydrophobic 1-benzylnicotinamide mimic, with which binding events were plausible both inside and outside the protein. This work demonstrates an extended substrate scope of the artificial metalloenzymes and provides information about the binding sites of the nicotinamide substrates, which can be exploited to further engineer artificial metalloenzymes for cofactor regeneration. Synopsis about graphical abstract: The manuscript provides information on the design of artificial metalloenzymes based on the bioconjugation of rhodium complexes to alcohol dehydrogenase, to improve their ability to reduce hydrophobic substrates. The graphical abstract presents different binding modes and results observed with native cofactors as substrates, compared to the hydrophobic benzylnicotinamide.
Citation
Basle, M., Padley, H. A., Martins, F. L., Winkler, G. S., Jäger, C. M., & Pordea, A. (2021). Design of artificial metalloenzymes for the reduction of nicotinamide cofactors. Journal of Inorganic Biochemistry, 220, Article 111446. https://doi.org/10.1016/j.jinorgbio.2021.111446
Journal Article Type | Article |
---|---|
Acceptance Date | Mar 24, 2021 |
Online Publication Date | Apr 2, 2021 |
Publication Date | 2021-07 |
Deposit Date | Jun 16, 2021 |
Publicly Available Date | Apr 3, 2022 |
Journal | Journal of Inorganic Biochemistry |
Print ISSN | 0162-0134 |
Electronic ISSN | 1873-3344 |
Publisher | Elsevier |
Peer Reviewed | Peer Reviewed |
Volume | 220 |
Article Number | 111446 |
DOI | https://doi.org/10.1016/j.jinorgbio.2021.111446 |
Public URL | https://nottingham-repository.worktribe.com/output/5490769 |
Publisher URL | https://www.sciencedirect.com/science/article/pii/S0162013421000933?via%3Dihub |
Files
JINORGBIO Accepted Manuscript
(829 Kb)
PDF
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