2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis
Abstract
Zymomonas mobilis is an industrially relevant bacterium notable for its ability to rapidly ferment simple sugars to ethanol using the Entner-Doudoroff (ED) glycolytic pathway, an alternative to the well-known Embden-Meyerhof-Parnas (EMP) pathway used by most organisms. Recent computational studies have predicted that the ED pathway is substantially more thermodynamically favorable than the EMP pathway, a potential factor explaining the high glycolytic rate in Z. mobilis. Here, to investigate the in vivo thermodynamics of the ED pathway and central carbon metabolism in Z. mobilis, we implemented a network-level approach that integrates quantitative metabolomics with 2H and 13C metabolic flux analysis to estimate reversibility and Gibbs free energy (ΔG) of metabolic reactions. This analysis revealed a strongly thermodynamically favorable ED pathway in Z. mobilis that is nearly twice as favorable as the EMP pathway in E. coli or S. cerevisiae. The in vivo step-by-step thermodynamic profile of the ED pathway was highly similar to previous in silico predictions, indicating that maximizing ΔG for each pathway step likely constitutes a cellular objective in Z. mobilis. Our analysis also revealed novel features of Z. mobilis metabolism, including phosphofructokinase-like enzyme activity, tricarboxylic acid cycle anaplerosis via PEP carboxylase, and a metabolic imbalance in the pentosemore »
- Authors:
- Publication Date:
- Research Org.:
- Univ. of Wisconsin Madison, Madison, WI (United States); Great Lakes Bioenergy Research Center, Madison WI (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division
- Contributing Org.:
- University of Wisconsin-Madison
- OSTI Identifier:
- 1547206
- Alternate Identifier(s):
- OSTI ID: 1526976
- Grant/Contract Number:
- SC0018409; FC02-07ER64494; AC05-00OR22725; 4000136894; SC0018998
- Resource Type:
- Published Article
- Journal Name:
- Metabolic Engineering
- Additional Journal Information:
- Journal Name: Metabolic Engineering Journal Volume: 54 Journal Issue: C; Journal ID: ISSN 1096-7176
- Publisher:
- Elsevier
- Country of Publication:
- Belgium
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; 09 BIOMASS FUELS; Zymomonas mobilis; biofuels; thermodynamics; glycolysis; Mass spectrometry; microbial metabolism; isotope tracers; MFA; Gibbs energy
Citation Formats
Jacobson, Tyler B., Adamczyk, Paul A., Stevenson, David M., Regner, Matthew, Ralph, John, Reed, Jennifer L., and Amador-Noguez, Daniel. 2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis. Belgium: N. p., 2019.
Web. doi:10.1016/j.ymben.2019.05.006.
Jacobson, Tyler B., Adamczyk, Paul A., Stevenson, David M., Regner, Matthew, Ralph, John, Reed, Jennifer L., & Amador-Noguez, Daniel. 2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis. Belgium. https://doi.org/10.1016/j.ymben.2019.05.006
Jacobson, Tyler B., Adamczyk, Paul A., Stevenson, David M., Regner, Matthew, Ralph, John, Reed, Jennifer L., and Amador-Noguez, Daniel. Mon .
"2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis". Belgium. https://doi.org/10.1016/j.ymben.2019.05.006.
@article{osti_1547206,
title = {2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis},
author = {Jacobson, Tyler B. and Adamczyk, Paul A. and Stevenson, David M. and Regner, Matthew and Ralph, John and Reed, Jennifer L. and Amador-Noguez, Daniel},
abstractNote = {Zymomonas mobilis is an industrially relevant bacterium notable for its ability to rapidly ferment simple sugars to ethanol using the Entner-Doudoroff (ED) glycolytic pathway, an alternative to the well-known Embden-Meyerhof-Parnas (EMP) pathway used by most organisms. Recent computational studies have predicted that the ED pathway is substantially more thermodynamically favorable than the EMP pathway, a potential factor explaining the high glycolytic rate in Z. mobilis. Here, to investigate the in vivo thermodynamics of the ED pathway and central carbon metabolism in Z. mobilis, we implemented a network-level approach that integrates quantitative metabolomics with 2H and 13C metabolic flux analysis to estimate reversibility and Gibbs free energy (ΔG) of metabolic reactions. This analysis revealed a strongly thermodynamically favorable ED pathway in Z. mobilis that is nearly twice as favorable as the EMP pathway in E. coli or S. cerevisiae. The in vivo step-by-step thermodynamic profile of the ED pathway was highly similar to previous in silico predictions, indicating that maximizing ΔG for each pathway step likely constitutes a cellular objective in Z. mobilis. Our analysis also revealed novel features of Z. mobilis metabolism, including phosphofructokinase-like enzyme activity, tricarboxylic acid cycle anaplerosis via PEP carboxylase, and a metabolic imbalance in the pentose phosphate pathway resulting in excretion of shikimate pathway intermediates. As a result, the integrated approach we present here for in vivo ΔG quantitation may be applied to the thermodynamic profiling of pathways and metabolic networks in other microorganisms and will contribute to the development of quantitative models of metabolism.},
doi = {10.1016/j.ymben.2019.05.006},
journal = {Metabolic Engineering},
number = C,
volume = 54,
place = {Belgium},
year = {Mon Jul 01 00:00:00 EDT 2019},
month = {Mon Jul 01 00:00:00 EDT 2019}
}
https://doi.org/10.1016/j.ymben.2019.05.006
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