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Title: Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus

Abstract

Here, the anaerobic metabolism of crotonate, benzoate, and cyclohexane carboxylate by Syntrophus aciditrophicus grown syntrophically with Methanospirillum hungatei provides a model to study syntrophic cooperation. Recent studies revealed that S. aciditrophicus contains Re-citrate synthase but lacks the common Si-citrate synthase. To establish whether the Re-citrate synthase is involved in glutamate synthesis via the oxidative branch of the Krebs cycle, we have used [1-13C]acetate and [1-14C]acetate as well as [13C]bicarbonate as additional carbon sources during axenic growth of S. aciditrophicus on crotonate. Our analyses showed that labeled carbons were detected in at least 14 amino acids, indicating the global utilization of acetate and bicarbonate. The labeling patterns of alanine and aspartate verified that pyruvate and oxaloacetate were synthesized by consecutive carboxylations of acetyl coenzyme A (acetyl-CoA). The isotopomer profile and 13C nuclear magnetic resonance (NMR) spectroscopy of the obtained [13C]glutamate, as well as decarboxylation of [14C]glutamate, revealed that this amino acid was synthesized by two pathways. Unexpectedly, only the minor route used Re-citrate synthase (30 to 40%), whereas the majority of glutamate was synthesized via the reductive carboxylation of succinate. This symmetrical intermediate could have been formed from two acetates via hydration of crotonyl-CoA to 4-hydroxybutyryl-CoA. 4-Hydroxybutyrate was detected in themore » medium of S. aciditrophicus when grown on crotonate, but an active hydratase could not be measured in cell extracts, and the annotated 4-hydroxybutyryl-CoA dehydratase (SYN_02445) lacks key amino acids needed to catalyze the hydration of crotonyl-CoA. Besides Clostridium kluyveri, this study reveals the second example of a microbial species to employ two pathways for glutamate synthesis.« less

Authors:
 [1];  [2];  [3];  [4];  [4];  [2];  [2];  [5]
  1. Max Planck Inst. fur terrestrische Mikrobiologie, Marburg (Germany)
  2. Fachbereich Biologie and Synmikro, Philipps-Universität, Marburg, (Germany)
  3. Fachbereich Chemie, Philipps-Universität, Marburg (Germany)
  4. Washington Univ., St. Louis, MO (United States)
  5. Max Planck Inst. fur terrestrische Mikrobiologie, Marburg (Germany); Fachbereich Biologie and Synmikro, Philipps-Universität, Marburg, (Germany)
Publication Date:
Research Org.:
Washington Univ., St. Louis, MO (United States); Univ. of Oklahoma, Norman, OK (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22). Chemical Sciences, Geosciences & Biosciences Division
OSTI Identifier:
1470219
Grant/Contract Number:  
FG02-96ER20214
Resource Type:
Accepted Manuscript
Journal Name:
Applied and Environmental Microbiology
Additional Journal Information:
Journal Volume: 81; Journal Issue: 24; Journal ID: ISSN 0099-2240
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 54 ENVIRONMENTAL SCIENCES; syntrophy; bioenergy; methanogenesis; glutamate

Citation Formats

Kim, Marie, Le, Huynh M., Xie, Xiulan, Feng, Xueyang, Tang, Yinjie J., Mouttaki, Housna, McInerney, Michael J., and Buckel, Wolfgang. Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus. United States: N. p., 2015. Web. doi:10.1128/AEM.02323-15.
Kim, Marie, Le, Huynh M., Xie, Xiulan, Feng, Xueyang, Tang, Yinjie J., Mouttaki, Housna, McInerney, Michael J., & Buckel, Wolfgang. Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus. United States. doi:10.1128/AEM.02323-15.
Kim, Marie, Le, Huynh M., Xie, Xiulan, Feng, Xueyang, Tang, Yinjie J., Mouttaki, Housna, McInerney, Michael J., and Buckel, Wolfgang. Fri . "Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus". United States. doi:10.1128/AEM.02323-15. https://www.osti.gov/servlets/purl/1470219.
@article{osti_1470219,
title = {Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus},
author = {Kim, Marie and Le, Huynh M. and Xie, Xiulan and Feng, Xueyang and Tang, Yinjie J. and Mouttaki, Housna and McInerney, Michael J. and Buckel, Wolfgang},
abstractNote = {Here, the anaerobic metabolism of crotonate, benzoate, and cyclohexane carboxylate by Syntrophus aciditrophicus grown syntrophically with Methanospirillum hungatei provides a model to study syntrophic cooperation. Recent studies revealed that S. aciditrophicus contains Re-citrate synthase but lacks the common Si-citrate synthase. To establish whether the Re-citrate synthase is involved in glutamate synthesis via the oxidative branch of the Krebs cycle, we have used [1-13C]acetate and [1-14C]acetate as well as [13C]bicarbonate as additional carbon sources during axenic growth of S. aciditrophicus on crotonate. Our analyses showed that labeled carbons were detected in at least 14 amino acids, indicating the global utilization of acetate and bicarbonate. The labeling patterns of alanine and aspartate verified that pyruvate and oxaloacetate were synthesized by consecutive carboxylations of acetyl coenzyme A (acetyl-CoA). The isotopomer profile and 13C nuclear magnetic resonance (NMR) spectroscopy of the obtained [13C]glutamate, as well as decarboxylation of [14C]glutamate, revealed that this amino acid was synthesized by two pathways. Unexpectedly, only the minor route used Re-citrate synthase (30 to 40%), whereas the majority of glutamate was synthesized via the reductive carboxylation of succinate. This symmetrical intermediate could have been formed from two acetates via hydration of crotonyl-CoA to 4-hydroxybutyryl-CoA. 4-Hydroxybutyrate was detected in the medium of S. aciditrophicus when grown on crotonate, but an active hydratase could not be measured in cell extracts, and the annotated 4-hydroxybutyryl-CoA dehydratase (SYN_02445) lacks key amino acids needed to catalyze the hydration of crotonyl-CoA. Besides Clostridium kluyveri, this study reveals the second example of a microbial species to employ two pathways for glutamate synthesis.},
doi = {10.1128/AEM.02323-15},
journal = {Applied and Environmental Microbiology},
number = 24,
volume = 81,
place = {United States},
year = {2015},
month = {10}
}

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