Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes
Abstract
This study constructed cell‐free glycolytic enzyme systems and compared them to their in vivo functions in Escherichia coli . Under in vitro conditions, flux regulation followed enzyme concentrations and kinetics. In E. coli, only one of the isozymes of phosphofructokinase (PfkA) and fructose‐bisphosphate aldolase (FbaA) facilitate Embden‐Meyerhof‐Parnas (EMP) flux, but under in vitro assays, these isozymes were interchangeable. Additionally, in vitro introduction of the Entner–Doudoroff (ED) pathway improved glycolysis rates, while in vivo overexpression of the ED pathway could not capture significant flux unless its phosphotransferase system (PTS) was knocked out. Lastly, in vivo dynamic 13 C‐experiments revealed that the labeling order of EMP pathway intermediates was not strictly cascade, indicating intracellular metabolites were not well mixed. These enigmatic observations cannot be fully explained by thermodynamics or substrate level regulations. This article supports the long‐time conjecture that EMP enzymes are channeled, and the PTS may be an anchor point to initiate enzyme assemblies. © 2018 American Institute of Chemical Engineers AIChE J , 65: 483–490, 2019
- Authors:
-
- Dept. of Energy, Environmental and Chemical Engineering Washington University St. Louis MO 63130
- Key Laboratory of Combinatorial Biosynthesis and Drug Discovery Ministry of Education and Wuhan University School of Pharmaceutical Sciences Wuhan 430071 China
- Lawrence Berkeley National Laboratory Emeryville CA 64608
- Publication Date:
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 1472164
- Grant/Contract Number:
- DESC0018324
- Resource Type:
- Publisher's Accepted Manuscript
- Journal Name:
- AIChE Journal
- Additional Journal Information:
- Journal Name: AIChE Journal Journal Volume: 65 Journal Issue: 2; Journal ID: ISSN 0001-1541
- Publisher:
- Wiley Blackwell (John Wiley & Sons)
- Country of Publication:
- United States
- Language:
- English
Citation Formats
Abernathy, Mary H., Zhang, Yuchen, Hollinshead, Whitney D., Wang, George, Baidoo, Edward E. K., Liu, Tiangang, and Tang, Yinjie J. Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes. United States: N. p., 2018.
Web. doi:10.1002/aic.16367.
Abernathy, Mary H., Zhang, Yuchen, Hollinshead, Whitney D., Wang, George, Baidoo, Edward E. K., Liu, Tiangang, & Tang, Yinjie J. Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes. United States. https://doi.org/10.1002/aic.16367
Abernathy, Mary H., Zhang, Yuchen, Hollinshead, Whitney D., Wang, George, Baidoo, Edward E. K., Liu, Tiangang, and Tang, Yinjie J. Fri .
"Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes". United States. https://doi.org/10.1002/aic.16367.
@article{osti_1472164,
title = {Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes},
author = {Abernathy, Mary H. and Zhang, Yuchen and Hollinshead, Whitney D. and Wang, George and Baidoo, Edward E. K. and Liu, Tiangang and Tang, Yinjie J.},
abstractNote = {This study constructed cell‐free glycolytic enzyme systems and compared them to their in vivo functions in Escherichia coli . Under in vitro conditions, flux regulation followed enzyme concentrations and kinetics. In E. coli, only one of the isozymes of phosphofructokinase (PfkA) and fructose‐bisphosphate aldolase (FbaA) facilitate Embden‐Meyerhof‐Parnas (EMP) flux, but under in vitro assays, these isozymes were interchangeable. Additionally, in vitro introduction of the Entner–Doudoroff (ED) pathway improved glycolysis rates, while in vivo overexpression of the ED pathway could not capture significant flux unless its phosphotransferase system (PTS) was knocked out. Lastly, in vivo dynamic 13 C‐experiments revealed that the labeling order of EMP pathway intermediates was not strictly cascade, indicating intracellular metabolites were not well mixed. These enigmatic observations cannot be fully explained by thermodynamics or substrate level regulations. This article supports the long‐time conjecture that EMP enzymes are channeled, and the PTS may be an anchor point to initiate enzyme assemblies. © 2018 American Institute of Chemical Engineers AIChE J , 65: 483–490, 2019},
doi = {10.1002/aic.16367},
journal = {AIChE Journal},
number = 2,
volume = 65,
place = {United States},
year = {Fri Sep 21 00:00:00 EDT 2018},
month = {Fri Sep 21 00:00:00 EDT 2018}
}
https://doi.org/10.1002/aic.16367
Web of Science
Works referenced in this record:
High-throughput metabolic flux analysis based on gas chromatography–mass spectrometry derived 13C constraints
journal, February 2004
- Fischer, Eliane; Zamboni, Nicola; Sauer, Uwe
- Analytical Biochemistry, Vol. 325, Issue 2
Direct Evidence for Metabolon Formation and Substrate Channeling in Recombinant TCA Cycle Enzymes
journal, August 2016
- Bulutoglu, Beyza; Garcia, Kristen E.; Wu, Fei
- ACS Chemical Biology, Vol. 11, Issue 10
Channeling in native microbial pathways: Implications and challenges for metabolic engineering
journal, November 2017
- Abernathy, Mary H.; He, Lian; Tang, Yinjie J.
- Biotechnology Advances, Vol. 35, Issue 6
Effect of a pyruvate kinase ( pykF -gene) knockout mutation on the control of gene expression and metabolic fluxes in Escherichia coli
journal, June 2004
- Siddiquee, Khandaker Al Zaid; Arauzo-Bravo, Marcos J.; Shimizu, Kazuyuki
- FEMS Microbiology Letters, Vol. 235, Issue 1
Metabolic engineering and adaptive evolution of Escherichia coli KO11 for ethanol production through the Entner-Doudoroff and the pentose phosphate pathways : Engineering of E-D and PP pathways in KO11 pgi- knockout
journal, November 2016
- Huerta-Beristain, Gerardo; Cabrera-Ruiz, Rosina; Hernandez-Chavez, Georgina
- Journal of Chemical Technology & Biotechnology, Vol. 92, Issue 5
Combination of Entner-Doudoroff Pathway with MEP Increases Isoprene Production in Engineered Escherichia coli
journal, December 2013
- Liu, Huaiwei; Sun, Yuanzhang; Ramos, Kristine Rose M.
- PLoS ONE, Vol. 8, Issue 12
Glycolytic Enzymes Associate Dynamically with Mitochondria in Response to Respiratory Demand and Support Substrate Channeling
journal, November 2007
- Graham, James W. A.; Williams, Thomas C. R.; Morgan, Megan
- The Plant Cell, Vol. 19, Issue 11
Possible Pitfalls of Flux Calculations Based on 13C-Labeling
journal, April 2001
- van Winden, Wouter; Verheijen, Peter; Heijnen, Sef
- Metabolic Engineering, Vol. 3, Issue 2
eQuilibrator--the biochemical thermodynamics calculator
journal, November 2011
- Flamholz, A.; Noor, E.; Bar-Even, A.
- Nucleic Acids Research, Vol. 40, Issue D1
Modulation of the Major Paths of Carbon in Photorespiratory Mutants of Synechocystis
journal, January 2011
- Huege, Jan; Goetze, Jan; Schwarz, Doreen
- PLoS ONE, Vol. 6, Issue 1
The role of dynamic enzyme assemblies and substrate channelling in metabolic regulation
journal, May 2018
- Sweetlove, Lee J.; Fernie, Alisdair R.
- Nature Communications, Vol. 9, Issue 1
Large-scale in vivo flux analysis shows rigidity and suboptimal performance of Bacillus subtilis metabolism
journal, May 2005
- Fischer, Eliane; Sauer, Uwe
- Nature Genetics, Vol. 37, Issue 6
In Vitro Reconstitution and Optimization of the Entire Pathway to Convert Glucose into Fatty Acid
journal, January 2017
- Liu, Zheng; Zhang, Yuchen; Jia, Xiaoge
- ACS Synthetic Biology, Vol. 6, Issue 4
Interaction between phosphofructokinase and aldolase from Saccharomyces cerevisiae studied by aqueous two-phase partitioning
journal, February 2001
- Matic, Sandra; Widell, Susanne; Åkerlund, Hans-Erik
- Journal of Chromatography B: Biomedical Sciences and Applications, Vol. 751, Issue 2
Characterization of physiological responses to 22 gene knockouts in Escherichia coli central carbon metabolism
journal, September 2016
- Long, Christopher P.; Gonzalez, Jacqueline E.; Sandoval, Nicholas R.
- Metabolic Engineering, Vol. 37
Adaptation for fast growth on glucose by differential expression of central carbon metabolism and gal regulon genes in an Escherichia coli strain lacking the phosphoenolpyruvate:carbohydrate phosphotransferase system
journal, March 2005
- Flores, Noemí; Flores, Salvador; Escalante, Adelfo
- Metabolic Engineering, Vol. 7, Issue 2
Substrate-driven chemotactic assembly in an enzyme cascade
journal, December 2017
- Zhao, Xi; Palacci, Henri; Yadav, Vinita
- Nature Chemistry, Vol. 10, Issue 3
Global metabolic regulation analysis for Escherichia coli K12 based on protein expression by 2-dimensional electrophoresis and enzyme activity measurement
journal, January 2003
- Peng, L.; Shimizu, K.
- Applied Microbiology and Biotechnology, Vol. 61, Issue 2, p. 163-178
Metabolic flux analysis in a nonstationary system: Fed-batch fermentation of a high yielding strain of E. coli producing 1,3-propanediol
journal, May 2007
- Antoniewicz, M.; Kraynie, D.; Laffend, L.
- Metabolic Engineering, Vol. 9, Issue 3
A genome-scale Escherichia coli kinetic metabolic model k-ecoli457 satisfying flux data for multiple mutant strains
journal, December 2016
- Khodayari, Ali; Maranas, Costas D.
- Nature Communications, Vol. 7, Issue 1
Intrinsically disordered linkers determine the interplay between phase separation and gelation in multivalent proteins
journal, November 2017
- Harmon, Tyler S.; Holehouse, Alex S.; Rosen, Michael K.
- eLife, Vol. 6
Genetic Basis of Growth Adaptation of Escherichia coli after Deletion of pgi, a Major Metabolic Gene
journal, November 2010
- Charusanti, Pep; Conrad, Tom M.; Knight, Eric M.
- PLoS Genetics, Vol. 6, Issue 11
Analysis of Carbon Metabolism in Escherichia coli Strains with an Inactive Phosphotransferase System by 13C Labeling and NMR Spectroscopy
journal, April 2002
- Flores, S.; Gosset, G.; Flores, N.
- Metabolic Engineering, Vol. 4, Issue 2
Glycolytic strategy as a tradeoff between energy yield and protein cost
journal, April 2013
- Flamholz, A.; Noor, E.; Bar-Even, A.
- Proceedings of the National Academy of Sciences, Vol. 110, Issue 24
Mapping photoautotrophic metabolism with isotopically nonstationary 13C flux analysis
journal, November 2011
- Young, Jamey D.; Shastri, Avantika A.; Stephanopoulos, Gregory
- Metabolic Engineering, Vol. 13, Issue 6
Krebs Cycle Metabolon: Structural Evidence of Substrate Channeling Revealed by Cross-Linking and Mass Spectrometry
journal, December 2014
- Wu, Fei; Minteer, Shelley
- Angewandte Chemie International Edition, Vol. 54, Issue 6
Allosteric and non-allosteric E. coli phosphofructokinases: effects on growth
journal, April 1978
- Robinson, John P.; Fraenkel, D. G.
- Biochemical and Biophysical Research Communications, Vol. 81, Issue 3
Improved Production of l-Threonine in Escherichia coli by Use of a DNA Scaffold System
journal, November 2012
- Lee, Jun Hyoung; Jung, Suk-Chae; Bui, Le Minh
- Applied and Environmental Microbiology, Vol. 79, Issue 3
A Map of Protein-Metabolite Interactions Reveals Principles of Chemical Communication
journal, January 2018
- Piazza, Ilaria; Kochanowski, Karl; Cappelletti, Valentina
- Cell, Vol. 172, Issue 1-2
Isotopically nonstationary 13 C flux analysis of changes in Arabidopsis thaliana leaf metabolism due to high light acclimation
journal, November 2014
- Ma, Fangfang; Jazmin, Lara J.; Young, Jamey D.
- Proceedings of the National Academy of Sciences, Vol. 111, Issue 47
Quantifying E. coli Proteome and Transcriptome with Single-Molecule Sensitivity in Single Cells
journal, July 2010
- Taniguchi, Y.; Choi, P. J.; Li, G. -W.
- Science, Vol. 329, Issue 5991
Global Rebalancing of Cellular Resources by Pleiotropic Point Mutations Illustrates a Multi-scale Mechanism of Adaptive Evolution
journal, April 2016
- Utrilla, Jose; O’Brien, Edward J.; Chen, Ke
- Cell Systems, Vol. 2, Issue 4
Examining Escherichia coli glycolytic pathways, catabolite repression, and metabolite channeling using Δpfk mutants
journal, October 2016
- Hollinshead, Whitney D.; Rodriguez, Sarah; Martin, Hector Garcia
- Biotechnology for Biofuels, Vol. 9, Issue 1
WUFlux: an open-source platform for 13C metabolic flux analysis of bacterial metabolism
journal, November 2016
- He, Lian; Wu, Stephen G.; Zhang, Muhan
- BMC Bioinformatics, Vol. 17, Issue 1
Substrate channeling and enzyme complexes for biotechnological applications
journal, November 2011
- Zhang, Y. -H. Percival
- Biotechnology Advances, Vol. 29, Issue 6
Synthetic protein scaffolds provide modular control over metabolic flux
journal, August 2009
- Dueber, John E.; Wu, Gabriel C.; Malmirchegini, G. Reza
- Nature Biotechnology, Vol. 27, Issue 8, p. 753-759
Quantitative estimation of channeling from early glycolytic intermediates to CO2 in intact Escherichia coli: Quantitation of channeling in intact E. coli
journal, June 2005
- Shearer, Georgia; Lee, Jennifer C.; Koo, Jia-an
- FEBS Journal, Vol. 272, Issue 13
Genome-Scale Architecture of Small Molecule Regulatory Networks and the Fundamental Trade-Off between Regulation and Enzymatic Activity
journal, September 2017
- Reznik, Ed; Christodoulou, Dimitris; Goldford, Joshua E.
- Cell Reports, Vol. 20, Issue 11
Biomolecular condensates: organizers of cellular biochemistry
journal, February 2017
- Banani, Salman F.; Lee, Hyun O.; Hyman, Anthony A.
- Nature Reviews Molecular Cell Biology, Vol. 18, Issue 5
Pathway engineering for the production of aromatic compounds in Escherichia coli
journal, May 1996
- Floras, Noemí; Xiao, Jimmy; Berry, Alan
- Nature Biotechnology, Vol. 14, Issue 5
New tools for mass isotopomer data evaluation in13C flux analysis: Mass isotope correction, data consistency checking, and precursor relationships
journal, January 2004
- Wahl, S. Aljoscha; Dauner, Michael; Wiechert, Wolfgang
- Biotechnology and Bioengineering, Vol. 85, Issue 3
Absolute metabolite concentrations and implied enzyme active site occupancy in Escherichia coli
journal, June 2009
- Bennett, Bryson D.; Kimball, Elizabeth H.; Gao, Melissa
- Nature Chemical Biology, Vol. 5, Issue 8
Substrate channelling as an approach to cascade reactions
journal, March 2016
- Wheeldon, Ian; Minteer, Shelley D.; Banta, Scott
- Nature Chemistry, Vol. 8, Issue 4