Expression of Phosphofructokinase Is Not Sufficient to Enable Embden-Meyerhof-Parnas Glycolysis in Zymomonas mobilis ZM4
Abstract
Zymomonas mobilis is a bacterium that produces ethanol from glucose at up to 97% of theoretical efficiency on a carbon basis. One factor contributing to the high efficiency of ethanol production is that Z. mobilis has a low biomass yield. The low biomass yield may be caused partly by the low ATP yield of the Entner-Doudoroff (ED) glycolytic pathway used by Z. mobilis, which produces only one ATP per glucose consumed. To test the hypothesis that ATP yield limits biomass yield in Z. mobilis, we attempted to introduce the Embden-Meyerhof-Parnas (EMP) glycolytic pathway (with double the ATP yield) by expressing phosphofructokinase (Pfk I) from Escherichia coli. Expression of Pfk I caused growth inhibition and resulted in accumulation of mutations in the pfkA gene. Co-expression of additional EMP enzymes, fructose bisphosphate aldolase (Fba) and triose phosphate isomerase (Tpi), with Pfk I did not enable EMP flux, and resulted in production of glycerol as a side product. Further analysis indicated that heterologous reactions may have operated in the reverse direction because of native metabolite concentrations. This study reveals how the metabolomic context of a chassis organism influences the range of pathways that can be added by heterologous expression.
- Authors:
-
- Michigan State Univ., East Lansing, MI (United States)
- Univ. of Wisconsin, Madison, WI (United States)
- Publication Date:
- Research Org.:
- Great Lakes Bioenergy Research Center (GLBRC), Madison, WI (United States); USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1579344
- Grant/Contract Number:
- SC0018409; FC02-07ER64494; AC02-05CH11231
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Frontiers in Microbiology
- Additional Journal Information:
- Journal Volume: 10; Journal ID: ISSN 1664-302X
- Publisher:
- Frontiers Research Foundation
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; Zymomonas mobilis; glycolysis; Embden-Meyerhof-Parnas pathway; Entner-Doudoroff pathway; metabolic engineering
Citation Formats
Felczak, Magdalena M., Jacobson, Tyler B., Ong, Wai Kit, Amador-Noguez, Daniel, and TerAvest, Michaela A. Expression of Phosphofructokinase Is Not Sufficient to Enable Embden-Meyerhof-Parnas Glycolysis in Zymomonas mobilis ZM4. United States: N. p., 2019.
Web. doi:10.3389/fmicb.2019.02270.
Felczak, Magdalena M., Jacobson, Tyler B., Ong, Wai Kit, Amador-Noguez, Daniel, & TerAvest, Michaela A. Expression of Phosphofructokinase Is Not Sufficient to Enable Embden-Meyerhof-Parnas Glycolysis in Zymomonas mobilis ZM4. United States. https://doi.org/10.3389/fmicb.2019.02270
Felczak, Magdalena M., Jacobson, Tyler B., Ong, Wai Kit, Amador-Noguez, Daniel, and TerAvest, Michaela A. Fri .
"Expression of Phosphofructokinase Is Not Sufficient to Enable Embden-Meyerhof-Parnas Glycolysis in Zymomonas mobilis ZM4". United States. https://doi.org/10.3389/fmicb.2019.02270. https://www.osti.gov/servlets/purl/1579344.
@article{osti_1579344,
title = {Expression of Phosphofructokinase Is Not Sufficient to Enable Embden-Meyerhof-Parnas Glycolysis in Zymomonas mobilis ZM4},
author = {Felczak, Magdalena M. and Jacobson, Tyler B. and Ong, Wai Kit and Amador-Noguez, Daniel and TerAvest, Michaela A.},
abstractNote = {Zymomonas mobilis is a bacterium that produces ethanol from glucose at up to 97% of theoretical efficiency on a carbon basis. One factor contributing to the high efficiency of ethanol production is that Z. mobilis has a low biomass yield. The low biomass yield may be caused partly by the low ATP yield of the Entner-Doudoroff (ED) glycolytic pathway used by Z. mobilis, which produces only one ATP per glucose consumed. To test the hypothesis that ATP yield limits biomass yield in Z. mobilis, we attempted to introduce the Embden-Meyerhof-Parnas (EMP) glycolytic pathway (with double the ATP yield) by expressing phosphofructokinase (Pfk I) from Escherichia coli. Expression of Pfk I caused growth inhibition and resulted in accumulation of mutations in the pfkA gene. Co-expression of additional EMP enzymes, fructose bisphosphate aldolase (Fba) and triose phosphate isomerase (Tpi), with Pfk I did not enable EMP flux, and resulted in production of glycerol as a side product. Further analysis indicated that heterologous reactions may have operated in the reverse direction because of native metabolite concentrations. This study reveals how the metabolomic context of a chassis organism influences the range of pathways that can be added by heterologous expression.},
doi = {10.3389/fmicb.2019.02270},
journal = {Frontiers in Microbiology},
number = ,
volume = 10,
place = {United States},
year = {Fri Sep 27 00:00:00 EDT 2019},
month = {Fri Sep 27 00:00:00 EDT 2019}
}
Web of Science
Works referenced in this record:
The Entner-Doudoroff pathway: history, physiology and molecular biology
journal, September 1992
- Conway, Tyrrell
- FEMS Microbiology Letters, Vol. 103, Issue 1
The Entner-Doudoroff pathway: history, physiology and molecular biology
journal, September 1992
- Conway, Tyrrell
- FEMS Microbiology Letters, Vol. 103, Issue 1
The route of ethanol formation in Zymomonas mobilis
journal, March 1966
- Dawes, Ea; Ribbons, Dw; Large, Pj
- Biochemical Journal, Vol. 98, Issue 3
The route of ethanol formation in Zymomonas mobilis
journal, March 1966
- Dawes, Ea; Ribbons, Dw; Large, Pj
- Biochemical Journal, Vol. 98, Issue 3
eQuilibrator--the biochemical thermodynamics calculator
journal, November 2011
- Flamholz, A.; Noor, E.; Bar-Even, A.
- Nucleic Acids Research, Vol. 40, Issue D1
NADH dehydrogenase deficiency results in low respiration rate and improved aerobic growth of Zymomonas mobilis
journal, March 2008
- Kalnenieks, U.; Galinina, N.; Strazdina, I.
- Microbiology, Vol. 154, Issue 3
Structure of the Zymomonas mobilis respiratory chain: oxygen affinity of electron transport and the role of cytochrome c peroxidase
journal, June 2014
- Balodite, E.; Strazdina, I.; Galinina, N.
- Microbiology, Vol. 160, Issue Pt_9
2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis
journal, July 2019
- Jacobson, Tyler B.; Adamczyk, Paul A.; Stevenson, David M.
- Metabolic Engineering, Vol. 54
Impact of Expression of EMP Enzymes on Glucose Metabolism in Zymomonas mobilis
journal, April 2013
- Chen, Rachel Ruizhen; Agrawal, Manoj; Mao, Zichao
- Applied Biochemistry and Biotechnology, Vol. 170, Issue 4
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
Metabolite concentrations, fluxes and free energies imply efficient enzyme usage
journal, May 2016
- Park, Junyoung O.; Rubin, Sara A.; Xu, Yi-Fan
- Nature Chemical Biology, Vol. 12, Issue 7
Metabolite concentrations, fluxes and free energies imply efficient enzyme usage
journal, May 2016
- Park, Junyoung O.; Rubin, Sara A.; Xu, Yi-Fan
- Nature Chemical Biology, Vol. 12, Issue 7
Rethinking glycolysis: on the biochemical logic of metabolic pathways
journal, May 2012
- Bar-Even, Arren; Flamholz, Avi; Noor, Elad
- Nature Chemical Biology, Vol. 8, Issue 6
Kinetics of alcohol production by zymomonas mobilis at high sugar concentrations
journal, April 1979
- Rogers, P. L.; Lee, Kye Joon; Tribe, D. E.
- Biotechnology Letters, Vol. 1, Issue 4
Cell Aggregation and Aerobic Respiration Are Important for Zymomonas mobilis ZM4 Survival in an Aerobic Minimal Medium
journal, March 2019
- Jones-Burrage, Sara E.; Kremer, Timothy A.; McKinlay, James B.
- Applied and Environmental Microbiology, Vol. 85, Issue 10
Growthcurver: an R package for obtaining interpretable metrics from microbial growth curves
journal, April 2016
- Sprouffske, Kathleen; Wagner, Andreas
- BMC Bioinformatics, Vol. 17, Issue 1
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
Functional Analysis, Overexpression, and Kinetic Characterization of Pyruvate Kinase from Methicillin-Resistant Staphylococcus aureus
journal, September 2010
- Zoraghi, Roya; See, Raymond H.; Gong, Huansheng
- Biochemistry, Vol. 49, Issue 35
Effect of oxygen on the metabolism of Zymomonas mobilis
journal, November 1984
- Bringer, Stephanie; Finn, Robert K.; Sahm, Hermann
- Archives of Microbiology, Vol. 139, Issue 4
OptSSeq explores enzyme expression and function landscapes to maximize isobutanol production rate
journal, March 2019
- Ghosh, Indro N.; Martien, Julia; Hebert, Alexander S.
- Metabolic Engineering, Vol. 52
Electron transport and oxidative stress in Zymomonas mobilis respiratory mutants
journal, January 2012
- Strazdina, Inese; Kravale, Zane; Galinina, Nina
- Archives of Microbiology, Vol. 194, Issue 6
Rethinking glycolysis: on the biochemical logic of metabolic pathways
journal, May 2012
- Bar-Even, Arren; Flamholz, Avi; Noor, Elad
- Nature Chemical Biology, Vol. 8, Issue 6
Fermentation of dihydroxyacetone by engineered Escherichia coli and Klebsiella variicola to products
journal, April 2018
- Wang, Liang; Chauliac, Diane; Rhee, Mun Su
- Proceedings of the National Academy of Sciences, Vol. 115, Issue 17
Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes
journal, September 2018
- Abernathy, Mary H.; Zhang, Yuchen; Hollinshead, Whitney D.
- AIChE Journal, Vol. 65, Issue 2
Functional Analysis, Overexpression, and Kinetic Characterization of Pyruvate Kinase from Methicillin-Resistant Staphylococcus aureus
journal, September 2010
- Zoraghi, Roya; See, Raymond H.; Gong, Huansheng
- Biochemistry, Vol. 49, Issue 35
Ethanol production by Zymomonas and Saccharomyces, advantages and disadvantages
journal, November 1983
- Karsch, Thomas; Stahl, Ulf; Esser, Karl
- European Journal of Applied Microbiology and Biotechnology, Vol. 18, Issue 6
NADH dehydrogenase deficiency results in low respiration rate and improved aerobic growth of Zymomonas mobilis
journal, March 2008
- Kalnenieks, U.; Galinina, N.; Strazdina, I.
- Microbiology, Vol. 154, Issue 3
Comparative studies of glycolytic pathways and channeling under in vitro and in vivo modes
journal, September 2018
- Abernathy, Mary H.; Zhang, Yuchen; Hollinshead, Whitney D.
- AIChE Journal, Vol. 65, Issue 2
Structure of the Zymomonas mobilis respiratory chain: oxygen affinity of electron transport and the role of cytochrome c peroxidase
journal, June 2014
- Balodite, E.; Strazdina, I.; Galinina, N.
- Microbiology, Vol. 160, Issue Pt_9
Electron transport and oxidative stress in Zymomonas mobilis respiratory mutants
journal, January 2012
- Strazdina, Inese; Kravale, Zane; Galinina, Nina
- Archives of Microbiology, Vol. 194, Issue 6
Growthcurver: an R package for obtaining interpretable metrics from microbial growth curves
journal, April 2016
- Sprouffske, Kathleen; Wagner, Andreas
- BMC Bioinformatics, Vol. 17, Issue 1
Influence of Aeration and of Pantothenate on Growth Yields of Zymomonas mobilis
journal, January 1965
- Belaïch, Jean-Pierre; Senez, Jacques C.
- Journal of Bacteriology, Vol. 89, Issue 5