Ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation [Identification of a ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation]
Abstract
Ferredoxin:NAD+ oxidoreductase (NADH-FNOR) catalyzes the transfer of electrons from reduced ferredoxin to NAD+. This enzyme has been hypothesized to be the main enzyme responsible for ferredoxin oxidization in the NADH-based ethanol pathway in Thermoanaerobacterium saccharolyticum; however, the corresponding gene has not yet been identified. Here, we identified the Tsac_1705 protein as a candidate FNOR based on the homology of its functional domains. We then confirmed its activity in vitro with a ferredoxin-based FNOR assay. To determine its role in metabolism, the tsac_1705 gene was deleted in different strains of T. saccharolyticum. In wild-type T. saccharolyticum, deletion of tsac_1705 resulted in a 75% loss of NADH-FNOR activity, which indicated that Tsac_1705 is the main NADH-FNOR in T. saccharolyticum. When both NADH- and NADPH-linked FNOR genes were deleted, the ethanol titer decreased and the ratio of ethanol to acetate approached unity, indicative of the absence of FNOR activity. As a result, we tested the effect of heterologous expression of Tsac_1705 in Clostridium thermocellum and found improvements in both the titer and the yield of ethanol.
- Authors:
-
- Dartmouth College, Hanover, NH (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
- National Renewable Energy Lab. (NREL), Golden, CO (United States)
- Publication Date:
- Research Org.:
- National Renewable Energy Laboratory (NREL), Golden, CO (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1339505
- Report Number(s):
- NREL/JA-2700-67275
Journal ID: ISSN 0099-2240
- Grant/Contract Number:
- AC36-08GO28308
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Applied and Environmental Microbiology
- Additional Journal Information:
- Journal Volume: 82; Journal Issue: 24; Journal ID: ISSN 0099-2240
- Publisher:
- American Society for Microbiology
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 09 BIOMASS FUELS; 59 BASIC BIOLOGICAL SCIENCES; metabolic engineering; ferredoxins; ethanol production
Citation Formats
Tian, Liang, Lo, Jonathan, Shao, Xiongjun, Zheng, Tianyong, Olson, Daniel G., and Lynd, Lee R. Ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation [Identification of a ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation]. United States: N. p., 2016.
Web. doi:10.1128/AEM.02130-16.
Tian, Liang, Lo, Jonathan, Shao, Xiongjun, Zheng, Tianyong, Olson, Daniel G., & Lynd, Lee R. Ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation [Identification of a ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation]. United States. https://doi.org/10.1128/AEM.02130-16
Tian, Liang, Lo, Jonathan, Shao, Xiongjun, Zheng, Tianyong, Olson, Daniel G., and Lynd, Lee R. Fri .
"Ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation [Identification of a ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation]". United States. https://doi.org/10.1128/AEM.02130-16. https://www.osti.gov/servlets/purl/1339505.
@article{osti_1339505,
title = {Ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation [Identification of a ferredoxin:NAD+ oxidoreductase of Thermoanaerobacterium saccharolyticum and its role in ethanol formation]},
author = {Tian, Liang and Lo, Jonathan and Shao, Xiongjun and Zheng, Tianyong and Olson, Daniel G. and Lynd, Lee R.},
abstractNote = {Ferredoxin:NAD+ oxidoreductase (NADH-FNOR) catalyzes the transfer of electrons from reduced ferredoxin to NAD+. This enzyme has been hypothesized to be the main enzyme responsible for ferredoxin oxidization in the NADH-based ethanol pathway in Thermoanaerobacterium saccharolyticum; however, the corresponding gene has not yet been identified. Here, we identified the Tsac_1705 protein as a candidate FNOR based on the homology of its functional domains. We then confirmed its activity in vitro with a ferredoxin-based FNOR assay. To determine its role in metabolism, the tsac_1705 gene was deleted in different strains of T. saccharolyticum. In wild-type T. saccharolyticum, deletion of tsac_1705 resulted in a 75% loss of NADH-FNOR activity, which indicated that Tsac_1705 is the main NADH-FNOR in T. saccharolyticum. When both NADH- and NADPH-linked FNOR genes were deleted, the ethanol titer decreased and the ratio of ethanol to acetate approached unity, indicative of the absence of FNOR activity. As a result, we tested the effect of heterologous expression of Tsac_1705 in Clostridium thermocellum and found improvements in both the titer and the yield of ethanol.},
doi = {10.1128/AEM.02130-16},
journal = {Applied and Environmental Microbiology},
number = 24,
volume = 82,
place = {United States},
year = {Fri Sep 30 00:00:00 EDT 2016},
month = {Fri Sep 30 00:00:00 EDT 2016}
}
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