Uncovering rare NADH-preferring ketol-acid reductoisomerases
Abstract
All members of the ketol-acid reductoisomerase (KARI) enzyme family characterized to date have been shown to prefer the nicotinamide adenine dinucleotide phosphate hydride (NADPH) cofactor to nicotinamide adenine dinucleotide hydride (NADH). However, KARIs with the reversed cofactor preference are desirable for industrial applications, including anaerobic fermentation to produce branched-chain amino acids. By applying insights gained from structural and engineering studies of this enzyme family to a comprehensive multiple sequence alignment of KARIs, we identified putative NADH-utilizing KARIs and characterized eight whose catalytic efficiencies using NADH were equal to or greater than NADPH. These are the first naturally NADH-preferring KARIs reported and demonstrate that this property has evolved independently multiple times, using strategies unlike those used previously in the laboratory to engineer a KARI cofactor switch.
Additional Information
© 2014 Elsevier B.V. Received 20 June 2014, Revised 1 August 2014, Accepted 19 August 2014, Available online 27 August 2014. J.K.B.C. acknowledges the support of the Resnick Sustainability Institute (Caltech). This publication is funded by the Gordon and Betty Moore Foundation through Grant GBMF2809 to the Caltech Programmable Molecular Technology Initiative. Author contributions: SBC, JKBC, and FHA designed research. SBC and JKBC performed research and analyzed data. SBC, JKBC, and FHA wrote the paper.Attached Files
Accepted Version - JACKSON-1-s2.0-S1096717614001062-main.pdf
Supplemental Material - mmc1.doc
Supplemental Material - mmc2.doc
Files
Name | Size | Download all |
---|---|---|
md5:f13b7d156ce376b5c9d319e1826dc40f
|
52.2 kB | Download |
md5:8f57e1545b6f637a135b1173837cc811
|
98.9 kB | Download |
md5:4e1361ec844674f0973429ba445060ba
|
723.7 kB | Preview Download |
Additional details
- Eprint ID
- 49116
- Resolver ID
- CaltechAUTHORS:20140902-115405020
- Resnick Sustainability Institute
- Gordon and Betty Moore Foundation
- GBMF2809
- Caltech Programmable Molecular Technology Initiative
- Created
-
2014-09-02Created from EPrint's datestamp field
- Updated
-
2021-11-10Created from EPrint's last_modified field
- Caltech groups
- Resnick Sustainability Institute