skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum

Abstract

Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are prominent candidate biocatalysts that, together, can enable the direct biotic conversion of lignocellulosic biomass to ethanol. The imbalance and suboptimal turnover rates of redox cofactors are currently hindering engineering efforts to achieve higher bioproductivity in both organisms. Measuring relevant intracellular cofactor concentrations will help understand redox state of these cofactors and help identify a strategy to overcome these limitations; however, metabolomic determinations of these labile metabolites have historically proved challenging.Results: Through our validations, we verified the handling and storage stability of these metabolites, and verified extraction matrices and extraction solvent were not suppressing mass spectrometry signals. We recovered adenylate energy charge ratios (a main quality indicator) above 0.82 for all extractions. NADH/NAD+ values of 0.26 and 0.04 for an adhE-deficient strain of C. thermocellum and its parent, respectively, reflect the expected shift to a more reduced redox potential when a species lacks the ability to re-oxidize NADH by synthesizing ethanol. This method failed to yield reliable results with C. bescii and poor-growing strains of T. saccharolyticum. Lastly, our validated protocols demonstrate and validate the extraction and analysis of selected redox and energy-related metabolites from two candidate consolidated bioprocessing biocatalysts, C. thermocellum and T. saccharolyticum.more » This development and validation highlights the important, but often neglected, need to optimize and validate metabolomic protocols when adapting them to new cell or tissue types.« less

Authors:
 [1]; ORCiD logo [2];  [3]; ORCiD logo [2]; ORCiD logo [4]; ORCiD logo [1]; ORCiD logo [2]
  1. Univ. of Tennessee, Knoxville, TN (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Centers for Disease Control and Prevention, Atlanta, GA (United States)
  4. Univ. of Tennessee, Knoxville, TN (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); LanzaTech, Skokie, IL (United States)
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1435314
Grant/Contract Number:
AC05-00OR22725
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Biotechnology for Biofuels
Additional Journal Information:
Journal Volume: 10; Journal Issue: 1; Journal ID: ISSN 1754-6834
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; Clostridium thermocellum; Thermoanaerobacterium saccharolyticum; Redox; Adenylate energy charge; Targeted metabolomics

Citation Formats

Sander, Kyle, Asano, Keiji G., Bhandari, Deepak, Van Berkel, Gary J., Brown, Steven D., Davison, Brian H., and Tschaplinski, Timothy J.. Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum. United States: N. p., 2017. Web. doi:10.1186/s13068-017-0960-4.
Sander, Kyle, Asano, Keiji G., Bhandari, Deepak, Van Berkel, Gary J., Brown, Steven D., Davison, Brian H., & Tschaplinski, Timothy J.. Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum. United States. doi:10.1186/s13068-017-0960-4.
Sander, Kyle, Asano, Keiji G., Bhandari, Deepak, Van Berkel, Gary J., Brown, Steven D., Davison, Brian H., and Tschaplinski, Timothy J.. Thu . "Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum". United States. doi:10.1186/s13068-017-0960-4. https://www.osti.gov/servlets/purl/1435314.
@article{osti_1435314,
title = {Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum},
author = {Sander, Kyle and Asano, Keiji G. and Bhandari, Deepak and Van Berkel, Gary J. and Brown, Steven D. and Davison, Brian H. and Tschaplinski, Timothy J.},
abstractNote = {Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are prominent candidate biocatalysts that, together, can enable the direct biotic conversion of lignocellulosic biomass to ethanol. The imbalance and suboptimal turnover rates of redox cofactors are currently hindering engineering efforts to achieve higher bioproductivity in both organisms. Measuring relevant intracellular cofactor concentrations will help understand redox state of these cofactors and help identify a strategy to overcome these limitations; however, metabolomic determinations of these labile metabolites have historically proved challenging.Results: Through our validations, we verified the handling and storage stability of these metabolites, and verified extraction matrices and extraction solvent were not suppressing mass spectrometry signals. We recovered adenylate energy charge ratios (a main quality indicator) above 0.82 for all extractions. NADH/NAD+ values of 0.26 and 0.04 for an adhE-deficient strain of C. thermocellum and its parent, respectively, reflect the expected shift to a more reduced redox potential when a species lacks the ability to re-oxidize NADH by synthesizing ethanol. This method failed to yield reliable results with C. bescii and poor-growing strains of T. saccharolyticum. Lastly, our validated protocols demonstrate and validate the extraction and analysis of selected redox and energy-related metabolites from two candidate consolidated bioprocessing biocatalysts, C. thermocellum and T. saccharolyticum. This development and validation highlights the important, but often neglected, need to optimize and validate metabolomic protocols when adapting them to new cell or tissue types.},
doi = {10.1186/s13068-017-0960-4},
journal = {Biotechnology for Biofuels},
number = 1,
volume = 10,
place = {United States},
year = {Thu Nov 30 00:00:00 EST 2017},
month = {Thu Nov 30 00:00:00 EST 2017}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Save / Share: