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Title: Flux analysis of central metabolic pathways in Geobactermetallireducens during reduction of solubleFe(III)-NTA

Abstract

We analyzed the carbon fluxes in the central metabolism ofGeobacter metallireducens strain GS-15 using 13C isotopomer modeling.Acetate labeled in the 1st or 2nd position was the sole carbon source,and Fe-NTA was the sole terminal electron acceptor. The measured labeledacetate uptake rate was 21 mmol/gdw/h in the exponential growth phase.The resulting isotope labeling pattern of amino acids allowed an accuratedetermination of the in vivo global metabolic reaction rates (fluxes)through the central metabolic pathways using a computational isotopomermodel. The tracer experiments showed that G. metallireducens containedcomplete biosynthesis pathways for essential metabolism, and this strainmight also have an unusual isoleucine biosynthesis route (usingacetyl-CoA and pyruvate as the precursors). The model indicated that over90 percent of the acetate was completely oxidized to CO2 via a completetricarboxylic acid (TCA) cycle while reducing iron. Pyruvate carboxylaseand phosphoenolpyruvate carboxykinase were present under theseconditions, but enzymes in the glyoxylate shunt and malic enzyme wereabsent. Gluconeogenesis and the pentose phosphate pathway were mainlyemployed for biosynthesis and accounted for less than 3 percent of totalcarbon consumption. The model also indicated surprisingly highreversibility in the reaction between oxoglutarate and succinate. Thisstep operates close to the thermodynamic equilibrium possibly becausesuccinate is synthesized via a transferase reaction, and the conversionof oxoglutarate to succinatemore » is a rate limiting step for carbonmetabolism. These findings enable a better understanding of therelationship between genome annotation and extant metabolic pathways inG. metallireducens.« less

Authors:
; ; ; ; ;
Publication Date:
Research Org.:
Ernest Orlando Lawrence Berkeley NationalLaboratory, Berkeley, CA (US)
Sponsoring Org.:
USDOE Director. Office of Science. Biological andEnvironmental Research
OSTI Identifier:
918684
Report Number(s):
LBNL-63533
Journal ID: ISSN 0099-2240; AEMIDF; R&D Project: VGTLTH; BnR: KP1102010; TRN: US200819%%405
DOE Contract Number:
DE-AC02-05CH11231
Resource Type:
Journal Article
Resource Relation:
Journal Name: Applied and Environmental Microbiology; Journal Volume: 73; Journal Issue: 12; Related Information: Journal Publication Date: June 2007
Country of Publication:
United States
Language:
English
Subject:
54 ENVIRONMENTAL SCIENCES; AMINO ACIDS; BINDING ENERGY; BIOLOGICAL PATHWAYS; BIOSYNTHESIS; CARBON; CARBON SOURCES; IN VIVO; METABOLISM; REACTION KINETICS; labeled acetate minimal medium isoleucine TCA cycle acetyl-CoAtransferase

Citation Formats

Tang, Yinjie J., Chakraborty, Romy, Garcia-Martin, Hector, Chu,Jeannie, Hazen, Terry C., and Keasling, Jay D. Flux analysis of central metabolic pathways in Geobactermetallireducens during reduction of solubleFe(III)-NTA. United States: N. p., 2007. Web. doi:10.1128/AEM.02986-06.
Tang, Yinjie J., Chakraborty, Romy, Garcia-Martin, Hector, Chu,Jeannie, Hazen, Terry C., & Keasling, Jay D. Flux analysis of central metabolic pathways in Geobactermetallireducens during reduction of solubleFe(III)-NTA. United States. doi:10.1128/AEM.02986-06.
Tang, Yinjie J., Chakraborty, Romy, Garcia-Martin, Hector, Chu,Jeannie, Hazen, Terry C., and Keasling, Jay D. Mon . "Flux analysis of central metabolic pathways in Geobactermetallireducens during reduction of solubleFe(III)-NTA". United States. doi:10.1128/AEM.02986-06. https://www.osti.gov/servlets/purl/918684.
@article{osti_918684,
title = {Flux analysis of central metabolic pathways in Geobactermetallireducens during reduction of solubleFe(III)-NTA},
author = {Tang, Yinjie J. and Chakraborty, Romy and Garcia-Martin, Hector and Chu,Jeannie and Hazen, Terry C. and Keasling, Jay D.},
abstractNote = {We analyzed the carbon fluxes in the central metabolism ofGeobacter metallireducens strain GS-15 using 13C isotopomer modeling.Acetate labeled in the 1st or 2nd position was the sole carbon source,and Fe-NTA was the sole terminal electron acceptor. The measured labeledacetate uptake rate was 21 mmol/gdw/h in the exponential growth phase.The resulting isotope labeling pattern of amino acids allowed an accuratedetermination of the in vivo global metabolic reaction rates (fluxes)through the central metabolic pathways using a computational isotopomermodel. The tracer experiments showed that G. metallireducens containedcomplete biosynthesis pathways for essential metabolism, and this strainmight also have an unusual isoleucine biosynthesis route (usingacetyl-CoA and pyruvate as the precursors). The model indicated that over90 percent of the acetate was completely oxidized to CO2 via a completetricarboxylic acid (TCA) cycle while reducing iron. Pyruvate carboxylaseand phosphoenolpyruvate carboxykinase were present under theseconditions, but enzymes in the glyoxylate shunt and malic enzyme wereabsent. Gluconeogenesis and the pentose phosphate pathway were mainlyemployed for biosynthesis and accounted for less than 3 percent of totalcarbon consumption. The model also indicated surprisingly highreversibility in the reaction between oxoglutarate and succinate. Thisstep operates close to the thermodynamic equilibrium possibly becausesuccinate is synthesized via a transferase reaction, and the conversionof oxoglutarate to succinate is a rate limiting step for carbonmetabolism. These findings enable a better understanding of therelationship between genome annotation and extant metabolic pathways inG. metallireducens.},
doi = {10.1128/AEM.02986-06},
journal = {Applied and Environmental Microbiology},
number = 12,
volume = 73,
place = {United States},
year = {Mon Jan 01 00:00:00 EST 2007},
month = {Mon Jan 01 00:00:00 EST 2007}
}