Creation of Plasmidless and Markerless Escherichia coli Succinate Producing Strain and Evaluation of its Biosynthetic Potential during Utilization of Lignocellulosic Sugars

2020 ◽  
Vol 36 (2) ◽  
pp. 3-11
Author(s):  
O.A. Zhuravliova ◽  
Т.А. Voeikova ◽  
A.Yu. Gulevich ◽  
V.G. Debabov
Keyword(s):  
Escherichia Coli ◽  
Succinic Acid ◽  
Plant Biomass ◽  
Anaerobic Fermentation ◽  
Basic Research ◽  
Acid Fermentation ◽  
Mixed Acid ◽  
Gene Coding ◽  
Lignocellulosic Sugars ◽  
Basic Research Project

The plasmidless and markerless Escherichia coli succinate producing strain SGM2.0Pyc-int has been engineered and characterized. The strain has the inactivated main mixed-acid fermentation pathways due to the deletions of ldhA,poxB, ackA,pta, and adhE genes, constitutively expresses the genes of the aceEF-lpdA operon encoding components of pyravate dehydrogenase complex, and possesses the chromosomally integrated Bacillus subtilis pycA gene coding for pyruvate carboxylase. The capacity of the strain to synthesize succinic acid in course of dual-phase aerobic-anaerobic fermentation with lignocellulosic sugars as substrates was studied. The SGM2.0Pyc-int strain synthesized succinic acid from glucose, xylose, and arabinose with a molar yields of 1.41 mol/mol, 1.18 mol/mol, and 1.18 mol/mol, respectively, during the anaerobic production stage. The constructed strain has great potential for developing efficient processes for the succinic acid production from plant biomass-derived sugars. Escherichia coli, fermentation, arabinose, glucose, xylose, succinic acid. The work was supported by a Grant from the Russian Foundation for Basic Research (Project no. 18-29-14005).

scholarly journals Aerobic Fermentation of d-Glucose by an Evolved Cytochrome Oxidase-Deficient Escherichia coli Strain

2008 ◽  
Vol 74 (24) ◽  
pp. 7561-7569 ◽  
Author(s):  
Vasiliy A. Portnoy ◽  
Markus J. Herrgård ◽  
Bernhard Ø. Palsson
Keyword(s):  
Escherichia Coli ◽  
Oxygen Uptake ◽  
Growth Conditions ◽  
Aerobic Growth ◽  
Acid Fermentation ◽  
Mixed Acid Fermentation ◽  
E Coli ◽  
Knockout Strain ◽  
Mixed Acid ◽  
Cytochrome Oxidases

ABSTRACT Fermentation of glucose to d-lactic acid under aerobic growth conditions by an evolved Escherichia coli mutant deficient in three terminal oxidases is reported in this work. Cytochrome oxidases (cydAB, cyoABCD, and cbdAB) were removed from the E. coli K12 MG1655 genome, resulting in the ECOM3 (E. coli cytochrome oxidase mutant) strain. Removal of cytochrome oxidases reduced the oxygen uptake rate of the knockout strain by nearly 85%. Moreover, the knockout strain was initially incapable of growing on M9 minimal medium. After the ECOM3 strain was subjected to adaptive evolution on glucose M9 medium for 60 days, a growth rate equivalent to that of anaerobic wild-type E. coli was achieved. Our findings demonstrate that three independently adaptively evolved ECOM3 populations acquired different phenotypes: one produced lactate as a sole fermentation product, while the other two strains exhibited a mixed-acid fermentation under oxic growth conditions with lactate remaining as the major product. The homofermenting strain showed a d-lactate yield of 0.8 g/g from glucose. Gene expression and in silico model-based analyses were employed to identify perturbed pathways and explain phenotypic behavior. Significant upregulation of ygiN and sodAB explains the remaining oxygen uptake that was observed in evolved ECOM3 strains. E. coli strains produced in this study showed the ability to produce lactate as a fermentation product from glucose and to undergo mixed-acid fermentation during aerobic growth.

Engineering Escherichia coli with acrylate pathway genes for propionic acid synthesis and its impact on mixed-acid fermentation

2012 ◽  
Vol 97 (3) ◽  
pp. 1191-1200 ◽  
Author(s):  
Vijayalakshmi Kandasamy ◽  
Hema Vaidyanathan ◽  
Ivana Djurdjevic ◽  
Elamparithi Jayamani ◽  
K. B. Ramachandran ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Propionic Acid ◽  
Acid Synthesis ◽  
Acid Fermentation ◽  
Mixed Acid Fermentation ◽  
Mixed Acid ◽  
Acrylate Pathway

scholarly journals Harnessing Escherichia coli for bio-based production of formate under pressurized H2 and CO2 gases

2021 ◽  
Author(s):  
Magali Roger ◽  
Tom C. Reed ◽  
Frank Sargent
Keyword(s):  
Escherichia Coli ◽  
Carbon Dioxide ◽  
Carbon Capture ◽  
Carbon Capture And Storage ◽  
Host Strain ◽  
Acid Fermentation ◽  
E Coli ◽  
Mixed Acid ◽  
Batch Bioreactor ◽  
Growth Deficiency

ABSRACTEscherichia coli is gram-negative bacterium that is a workhorse of the biotechnology industry. The organism has a flexible metabolism and can perform a mixed-acid fermentation under anaerobic conditions. Under these conditions E. coli synthesises a formate hydrogenlyase isoenzyme (FHL-1) that can generate molecular hydrogen and carbon dioxide from formic acid. The reverse reaction is hydrogen-dependent carbon dioxide reduction (HDCR), which has exciting possibilities in bio-based carbon capture and storage if it can be harnessed. In this study, an E. coli host strain was optimised for the production of formate from H2 and CO2 during bacterial growth in a pressurised batch bioreactor. A host strain was engineered that constitutively produced the FHL-1 enzyme and incorporation of tungsten in to the enzyme, in place of molybdenum, helped poise the reaction in the HDCR direction. The engineered E. coli strain showed an ability to grow under fermentative conditions while simultaneously producing formate from gaseous H2 and CO2 supplied in the bioreactor. However, while a sustained pressure of 10 bar N2 had no adverse effect on cell growth, when the culture was placed at or above 4 bar pressure of a H2:CO2 mixture then a clear growth deficiency was observed. Taken together, this work demonstrates that growing cells can be harnessed to hydrogenate carbon dioxide and provides fresh evidence that the FHL-1 enzyme may be intimately linked with bacterial energy metabolism.

scholarly journals Metabolic engineering of the mixed-acid fermentation pathway of Escherichia coli for anaerobic production of glutamate and itaconate

AMB Express ◽  
2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Kiira S. Vuoristo ◽  
Astrid E. Mars ◽  
Jose Vidal Sangra ◽  
Jan Springer ◽  
Gerrit Eggink ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Metabolic Engineering ◽  
Acid Fermentation ◽  
Mixed Acid Fermentation ◽  
Mixed Acid ◽  
Anaerobic Production

Production of polyhydroxyalkanoates by Escherichia coli mutants with defected mixed acid fermentation pathways

2010 ◽  
Vol 87 (6) ◽  
pp. 2247-2256 ◽  
Author(s):  
Jia Jian ◽  
Shao-Qin Zhang ◽  
Zhen-Yu Shi ◽  
Wei Wang ◽  
Guo-Qiang Chen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Fermentation ◽  
Mixed Acid Fermentation ◽  
Mixed Acid

scholarly journals On-line analysis and in situ pH monitoring of mixed acid fermentation by Escherichia coli using combined FTIR and Raman techniques

2020 ◽  
Vol 412 (26) ◽  
pp. 7307-7319
Author(s):  
George D. Metcalfe ◽  
Thomas W. Smith ◽  
Michael Hippler
Keyword(s):  
Escherichia Coli ◽  
Detection Limits ◽  
Bacterial Suspension ◽  
Spectral Features ◽  
Order Kinetic ◽  
Acid Fermentation ◽  
Mixed Acid Fermentation ◽  
Mixed Acid ◽  
Formate Decomposition

Abstract We introduce an experimental setup allowing continuous monitoring of bacterial fermentation processes by simultaneous optical density (OD) measurements, long-path FTIR headspace monitoring of CO2, acetaldehyde and ethanol, and liquid Raman spectroscopy of acetate, formate, and phosphate anions, without sampling. We discuss which spectral features are best suited for detection, and how to obtain partial pressures and concentrations by integrations and least squares fitting of spectral features. Noise equivalent detection limits are about 2.6 mM for acetate and 3.6 mM for formate at 5 min integration time, improving to 0.75 mM for acetate and 1.0 mM for formate at 1 h integration. The analytical range extends to at least 1 M with a standard deviation of percentage error of about 8%. The measurement of the anions of the phosphate buffer allows the spectroscopic, in situ determination of the pH of the bacterial suspension via a modified Henderson-Hasselbalch equation in the 6–8 pH range with an accuracy better than 0.1. The 4 m White cell FTIR measurements provide noise equivalent detection limits of 0.21 μbar for acetaldehyde and 0.26 μbar for ethanol in the gas phase, corresponding to 3.2 μM acetaldehyde and 22 μM ethanol in solution, using Henry’s law. The analytical dynamic range exceeds 1 mbar ethanol corresponding to 85 mM in solution. As an application example, the mixed acid fermentation of Escherichia coli is studied. The production of CO2, ethanol, acetaldehyde, acids such as formate and acetate, and the changes in pH are discussed in the context of the mixed acid fermentation pathways. Formate decomposition into CO2 and H2 is found to be governed by a zeroth-order kinetic rate law, showing that adding exogenous formate to a bioreactor with E. coli is expected to have no beneficial effect on the rate of formate decomposition and biohydrogen production.

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