scholarly journals Systems-informed genome mining for electroautotrophic microbial production

2020 ◽  
Author(s):  
Anthony J. Abel ◽  
Jacob M. Hilzinger ◽  
Adam P. Arkin ◽  
Douglas S. Clark
Keyword(s):  
Carbon Fixation ◽  
Microbial Respiration ◽  
Tricarboxylic Acid Cycle ◽  
Genome Mining ◽  
Tricarboxylic Acid ◽  
Nitrate Respiration ◽  
Fixation Rate ◽  
Acid Cycle ◽  
Carbon Molecules ◽  
Reductive Tricarboxylic Acid Cycle

AbstractMicrobial electrosynthesis (MES) systems can store renewable energy and CO2 in many-carbon molecules inaccessible to abiotic electrochemistry. Here, we develop a multiphysics model to investigate the fundamental and practical limits of MES enabled by direct electron uptake and we identify organisms in which this biotechnological CO2-fixation strategy can be realized. Systematic model comparisons of microbial respiration and carbon fixation strategies revealed that, under aerobic conditions, the CO2 fixation rate is limited to <6 μmol/cm2/hr by O2 mass transport despite efficient electron utilization. In contrast, anaerobic nitrate respiration enables CO2 fixation rates >50 μmol/cm2/hr for microbes using the reductive tricarboxylic acid cycle. Phylogenetic analysis, validated by recapitulating experimental demonstrations of electroautotrophy, uncovered multiple probable electroautotrophic organisms and a significant number of genetically tractable strains that require heterologous expression of <5 proteins to gain electroautotrophic function. The model and analysis presented here will guide microbial engineering and reactor design for practical MES systems.

scholarly journals Evidence for Autotrophic CO2 Fixation via the Reductive Tricarboxylic Acid Cycle by Members of the ε Subdivision of Proteobacteria

2005 ◽  
Vol 187 (9) ◽  
pp. 3020-3027 ◽  
Author(s):  
Michael Hügler ◽  
Carl O. Wirsen ◽  
Georg Fuchs ◽  
Craig D. Taylor ◽  
Stefan M. Sievert
Keyword(s):  
Carbon Fixation ◽  
Tricarboxylic Acid Cycle ◽  
Calvin Cycle ◽  
Tricarboxylic Acid ◽  
Rrna Gene ◽  
Atp Citrate Lyase ◽  
Acid Cycle ◽  
Key Enzymes ◽  
Citrate Lyase ◽  
Reductive Tricarboxylic Acid Cycle

ABSTRACT Based on 16S rRNA gene surveys, bacteria of the ε subdivision of proteobacteria have been identified to be important members of microbial communities in a variety of environments, and quite a few have been demonstrated to grow autotrophically. However, no information exists on what pathway of autotrophic carbon fixation these bacteria might use. In this study, Thiomicrospira denitrificans and Candidatus Arcobacter sulfidicus, two chemolithoautotrophic sulfur oxidizers of the ε subdivision of proteobacteria, were examined for activities of the key enzymes of the known autotrophic CO2 fixation pathways. Both organisms contained activities of the key enzymes of the reductive tricarboxylic acid cycle, ATP citrate lyase, 2-oxoglutarate:ferredoxin oxidoreductase, and pyruvate:ferredoxin oxidoreductase. Furthermore, no activities of key enzymes of other CO2 fixation pathways, such as the Calvin cycle, the reductive acetyl coenzyme A pathway, and the 3-hydroxypropionate cycle, could be detected. In addition to the key enzymes, the activities of the other enzymes involved in the reductive tricarboxylic acid cycle could be measured. Sections of the genes encoding the α- and β-subunits of ATP citrate lyase could be amplified from both organisms. These findings represent the first direct evidence for the operation of the reductive tricarboxylic acid cycle for autotrophic CO2 fixation in ε-proteobacteria. Since ε-proteobacteria closely related to these two organisms are important in many habitats, such as hydrothermal vents, oxic-sulfidic interfaces, or oilfields, these results suggest that autotrophic CO2 fixation via the reductive tricarboxylic acid cycle might be more important than previously considered.

scholarly journals Growth Enhancement Facilitated by Gaseous CO2 through Heterologous Expression of Reductive Tricarboxylic acid Cycle Genes in Escherichia coli

Fermentation ◽  
2021 ◽  
Vol 7 (2) ◽  
pp. 98
Author(s):  
Shou-Chen Lo ◽  
En-Pei Isabel Chiang ◽  
Ya-Tang Yang ◽  
Si-Yu Li ◽  
Jian-Hau Peng ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Carbon Fixation ◽  
Tricarboxylic Acid Cycle ◽  
Anaerobic Respiration ◽  
Tricarboxylic Acid ◽  
Coli Strain ◽  
Growth Enhancement ◽  
Acid Cycle ◽  
E Coli ◽  
Reductive Tricarboxylic Acid Cycle

The enzymatic mechanisms of carbon fixation by autotrophs, such as the reductive tricarboxylic acid cycle (rTCA), have inspired biotechnological approaches to producing bio-based chemicals directly through CO2. To explore the possibility of constructing an rTCA cycle in Escherichia coli and to investigate their potential for CO2 assimilation, a total of ten genes encoding the key rTCA cycle enzymes, including α-ketoglutarate:ferredoxin oxidoreductase, ATP-dependent citrate lyase, and fumarate reductase/succinate dehydrogenase, were cloned into E. coli. The transgenic E. coli strain exhibited enhanced growth and the ability to assimilate external inorganic carbon with a gaseous CO2 supply. Further experiments conducted in sugar-free medium containing hydrogen as the electron donor and dimethyl sulfoxide (DMSO) as the electron acceptor proved that the strain is able to undergo anaerobic respiration, using CO2 as the major carbon source. The transgenic stain demonstrated CO2-enhanced growth, whereas the genes involved in chemotaxis, flagellar assembly, and acid-resistance were upregulated under the anaerobic respiration. Furthermore, metabolomic analysis demonstrated that the total concentrations of ATP, ADP, and AMP in the transgenic strain were higher than those in the vector control strain and these results coincided with the enhanced growth. Our approach offers a novel strategy to engineer E. coli for assimilating external gaseous CO2.

Evaluation of anaerobic glucose utilization by Escherichia coli strains with impaired fermentation ability upon respiration with external and internal electron acceptor

2020 ◽  
Vol 36 (3) ◽  
pp. 25-33
Author(s):  
A.Yu. Gulevich ◽  
V.G. Debabov
Keyword(s):  
Escherichia Coli ◽  
Electron Acceptor ◽  
Glucose Utilization ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Nitrate Respiration ◽  
Activity Levels ◽  
Ammonium Ions ◽  
Acid Fermentation ◽  
Acid Cycle

The characteristics of anaerobic glucose utilization and metabolite production by recombinant Escherichia coli strains with impaired fermentation ability upon respiration with pyruvate as an internal and nitrate as an external electron acceptor have been studied. It was found that respiration processes utilizing pyruvic acid as an endogenous electron acceptor and leading to the lactate and alanine formation were capable of mutual interference. After elimination of ammonium ions from the medium, the native activity levels of respiratory lactate dehydrogenases Did and LldD in E. coli strains deficient in the mixed acid fermentation pathways can almost completely compensate for the loss of activity of respiratory alanine dehydrogenase DadA, but are insufficient to maintain the entire intracellular redox balance. The addition of nitrate ions in the medium abolished alanine production by the strains despite the availability of ammonium ions, while the functionality of respiratory reduction of endogenous pyruvate to lactate retained in the studied strains even in the presence of a strong exogenous oxidant. Respiration with external electron acceptor provoked the activation of the oxidative tricarboxylic acid cycle in the strains. Anaerobic glucose utilization by the strain with interrupted tricarboxylic acid cycle increased during nitrate respiration, but remained restricted by the excessive generation of reducing equivalents in the residual reactions of the cycle. Escherichia coli, glucose, fermentation, respiration, pyruvate, lactate, alanine, nitrate, ammonium. The work was supported by a grant from the Russian Foundation for Basic Research (project #18-04-01222).

scholarly journals A Soluble NADH-Dependent Fumarate Reductase in the Reductive Tricarboxylic Acid Cycle of Hydrogenobacter thermophilus TK-6

2008 ◽  
Vol 190 (21) ◽  
pp. 7170-7177 ◽  
Author(s):  
Akane Miura ◽  
Masafumi Kameya ◽  
Hiroyuki Arai ◽  
Masaharu Ishii ◽  
Yasuo Igarashi
Keyword(s):  
Soluble Fraction ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Fumarate Reductase ◽  
High Sequence Identity ◽  
Catalytic Subunits ◽  
Acid Cycle ◽  
Genes Encoding ◽  
Hydrogenobacter Thermophilus ◽  
Reductive Tricarboxylic Acid Cycle

ABSTRACT Fumarate reductase (FRD) is an enzyme that reduces fumarate to succinate. In many organisms, it is bound to the membrane and uses electron donors such as quinol. In this study, an FRD from a thermophilic chemolithoautotrophic bacterium, Hydrogenobacter thermophilus TK-6, was purified and characterized. FRD activity using NADH as an electron donor was not detected in the membrane fraction but was found in the soluble fraction. The purified enzyme was demonstrated to be a novel type of FRD, consisting of five subunits. One subunit showed high sequence identity to the catalytic subunits of known FRDs. Although the genes of typical FRDs are assembled in a cluster, the five genes encoding the H. thermophilus FRD were distant from each other in the genome. Furthermore, phylogenetic analysis showed that the H. thermophilus FRD was located in a distinct position from those of known soluble FRDs. This is the first report of a soluble NADH-dependent FRD in Bacteria and of the purification of a FRD that operates in the reductive tricarboxylic acid cycle.

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