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Title: Glycolysis without pyruvate kinase in Clostridium thermocellum

The metabolism of Clostridium thermocellum is notable in that it assimilates sugar via the EMP pathway but does not possess a pyruvate kinase enzyme. In the wild type organism, there are three proposed pathways for conversion of phosphoenolpyruvate (PEP) to pyruvate, which differ in their cofactor usage. One path uses pyruvate phosphate dikinase (PPDK), another pathway uses the combined activities of PEP carboxykinase (PEPCK) and oxaloacetate decarboxylase (ODC). Yet another pathway, the malate shunt, uses the combined activities of PEPCK, malate dehydrogenase and malic enzyme. First we showed that there is no flux through the ODC pathway by enzyme assay. Flux through the remaining two pathways (PPDK and malate shunt) was determined by dynamic 13C labeling. In the wild-type strain, the malate shunt accounts for about 33 ± 2% of the flux to pyruvate, with the remainder via the PPDK pathway. Deletion of the ppdk gene resulted in a redirection of all pyruvate flux through the malate shunt. Lastly, this provides the first direct evidence of the in-vivo function of the malate shunt.
Authors:
 [1] ;  [2] ;  [2] ;  [1] ;  [1] ;  [1] ;  [3] ;  [1] ;  [2] ;  [1]
  1. Dartmouth College, Hanover, NH (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. ETH Zurich, Zurich (Switzerland)
  3. Univ. of Wisconsin-Madison, Madison, WI (United States)
Publication Date:
Grant/Contract Number:
AC05-00OR22725; AC02-05CH11231
Type:
Accepted Manuscript
Journal Name:
Metabolic Engineering
Additional Journal Information:
Journal Name: Metabolic Engineering; Journal ID: ISSN 1096-7176
Publisher:
Elsevier
Research Org:
Dartmouth College, Hanover, NH (United States)
Sponsoring Org:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; 13C flux analysis; pyruvate kinase; malate shunt; malic enzyme; malate dehydrogenase; oxaloacetate decarboxylase
OSTI Identifier:
1333931
Alternate Identifier(s):
OSTI ID: 1397061

Olson, Daniel G., Horl, Manuel, Fuhrer, Tobias, Cui, Jingxuan, Zhou, Jilai, Maloney, Marybeth I., Amador-Noguez, Daniel, Tian, Liang, Sauer, Uwe, and Lynd, Lee R.. Glycolysis without pyruvate kinase in Clostridium thermocellum. United States: N. p., Web. doi:10.1016/j.ymben.2016.11.011.
Olson, Daniel G., Horl, Manuel, Fuhrer, Tobias, Cui, Jingxuan, Zhou, Jilai, Maloney, Marybeth I., Amador-Noguez, Daniel, Tian, Liang, Sauer, Uwe, & Lynd, Lee R.. Glycolysis without pyruvate kinase in Clostridium thermocellum. United States. doi:10.1016/j.ymben.2016.11.011.
Olson, Daniel G., Horl, Manuel, Fuhrer, Tobias, Cui, Jingxuan, Zhou, Jilai, Maloney, Marybeth I., Amador-Noguez, Daniel, Tian, Liang, Sauer, Uwe, and Lynd, Lee R.. 2016. "Glycolysis without pyruvate kinase in Clostridium thermocellum". United States. doi:10.1016/j.ymben.2016.11.011. https://www.osti.gov/servlets/purl/1333931.
@article{osti_1333931,
title = {Glycolysis without pyruvate kinase in Clostridium thermocellum},
author = {Olson, Daniel G. and Horl, Manuel and Fuhrer, Tobias and Cui, Jingxuan and Zhou, Jilai and Maloney, Marybeth I. and Amador-Noguez, Daniel and Tian, Liang and Sauer, Uwe and Lynd, Lee R.},
abstractNote = {The metabolism of Clostridium thermocellum is notable in that it assimilates sugar via the EMP pathway but does not possess a pyruvate kinase enzyme. In the wild type organism, there are three proposed pathways for conversion of phosphoenolpyruvate (PEP) to pyruvate, which differ in their cofactor usage. One path uses pyruvate phosphate dikinase (PPDK), another pathway uses the combined activities of PEP carboxykinase (PEPCK) and oxaloacetate decarboxylase (ODC). Yet another pathway, the malate shunt, uses the combined activities of PEPCK, malate dehydrogenase and malic enzyme. First we showed that there is no flux through the ODC pathway by enzyme assay. Flux through the remaining two pathways (PPDK and malate shunt) was determined by dynamic 13C labeling. In the wild-type strain, the malate shunt accounts for about 33 ± 2% of the flux to pyruvate, with the remainder via the PPDK pathway. Deletion of the ppdk gene resulted in a redirection of all pyruvate flux through the malate shunt. Lastly, this provides the first direct evidence of the in-vivo function of the malate shunt.},
doi = {10.1016/j.ymben.2016.11.011},
journal = {Metabolic Engineering},
number = ,
volume = ,
place = {United States},
year = {2016},
month = {12}
}