scholarly journals Effect of deregulation of repressor-specific carbon catabolite repression on carbon source consumption in Escherichia coli

2021 ◽  
Vol 64 (1) ◽  
Author(s):  
Hyeon Jeong Seong ◽  
Yu-Sin Jang
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Catabolite Repression ◽  
Sole Carbon Source ◽  
Carbon Catabolite Repression ◽  
Cell Factory ◽  
Essential Information ◽  
E Coli ◽  
Marine Biomass ◽  
Galactose Utilization

AbstractEscherichia coli has been used as a host to construct the cell factory for biobased production of chemicals from renewable feedstocks. Because galactose is found in marine biomass as a major component, the strategy for galactose utilization in E. coli has been gained more attention. Although galactose and glucose co-fermentation has been reported using the engineered E. coli strain, few reports have covered fermentation supplemented with galactose as a sole carbon source in the mutant lacking the repressor-specific carbon catabolite repression (CCR). Here, we report the effects of the deregulation of the repressor-specific CCR (galR− and galS−) in fermentation supplemented with galactose as a sole carbon source, using the engineered E. coli strains. In the fermentation using the galR− and galS− double mutant (GR2 strain), an increase of rates in sugar consumption and cell growth was observed compared to the parent strain. In the glucose fermentation, wild-type W3110 and its mutant GR2 and GR2PZ (galR−, galS−, pfkA−, and zwf−) consumed sugar at a higher rate than those values obtained from galactose fermentation. However, the GR2P strain (galR−, galS−, and pfkA−) showed no difference between fermentations using glucose and galactose as a sole carbon source. This study provides essential information for galactose fermentation using the CCR-deregulated E. coli strains.

scholarly journals Heterologous Production of 6-Deoxyerythronolide B in Escherichia coli through the Wood Werkman Cycle

Metabolites ◽  
2020 ◽  
Vol 10 (6) ◽  
pp. 228
Author(s):  
R. Axayacatl Gonzalez-Garcia ◽  
Lars K. Nielsen ◽  
Esteban Marcellin
Keyword(s):  
Escherichia Coli ◽  
Natural Products ◽  
Carbon Source ◽  
Sole Carbon Source ◽  
Structural Diversity ◽  
Heterologous Production ◽  
E Coli ◽  
Anticancer Properties ◽  
Defined Media ◽  
Extender Unit

Polyketides are a remarkable class of natural products with diverse functional and structural diversity. The class includes many medicinally important molecules with antiviral, antimicrobial, antifungal and anticancer properties. Native bacterial, fungal and plant hosts are often difficult to cultivate and coax into producing the desired product. As a result, Escherichia coli has been used for the heterologous production of polyketides, with the production of 6-deoxyerythronolide B (6-dEB) being the first example. Current strategies for production in E. coli require feeding of exogenous propionate as a source for the precursors propionyl-CoA and S-methylmalonyl-CoA. Here, we show that heterologous polyketide production is possible from glucose as the sole carbon source. The heterologous expression of eight genes from the Wood-Werkman cycle found in Propionibacteria, in combination with expression of the 6-dEB synthases DEBS1, DEBS2 and DEBS3 resulted in 6-dEB formation from glucose as the sole carbon source. Our results show that the Wood-Werkman cycle provides the required propionyl-CoA and the extender unit S-methylmalonyl-CoA to produce up to 0.81 mg/L of 6-dEB in a chemically defined media.

scholarly journals Regulation of Arabinose and Xylose Metabolism in Escherichia coli

2009 ◽  
Vol 76 (5) ◽  
pp. 1524-1532 ◽  
Author(s):  
Tasha A. Desai ◽  
Christopher V. Rao
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Plant Biomass ◽  
Xylose Metabolism ◽  
Sugar Utilization ◽  
E Coli ◽  
Metabolic Genes ◽  
Pentose Sugars

ABSTRACT Bacteria such as Escherichia coli will often consume one sugar at a time when fed multiple sugars, in a process known as carbon catabolite repression. The classic example involves glucose and lactose, where E. coli will first consume glucose, and only when it has consumed all of the glucose will it begin to consume lactose. In addition to that of lactose, glucose also represses the consumption of many other sugars, including arabinose and xylose. In this work, we characterized a second hierarchy in E. coli, that between arabinose and xylose. We show that, when grown in a mixture of the two pentoses, E. coli will consume arabinose before it consumes xylose. Consistent with a mechanism involving catabolite repression, the expression of the xylose metabolic genes is repressed in the presence of arabinose. We found that this repression is AraC dependent and involves a mechanism where arabinose-bound AraC binds to the xylose promoters and represses gene expression. Collectively, these results demonstrate that sugar utilization in E. coli involves multiple layers of regulation, where cells will consume first glucose, then arabinose, and finally xylose. These results may be pertinent in the metabolic engineering of E. coli strains capable of producing chemical and biofuels from mixtures of hexose and pentose sugars derived from plant biomass.

Some biochemical effects of 4-deoxy-4-fluoro-D-glucose on Escherichia coli

1977 ◽  
Vol 55 (8) ◽  
pp. 911-915 ◽  
Author(s):  
N. F. Taylor ◽  
Li-Yu Louie
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Catabolite Repression ◽  
Dry Weight ◽  
Resting Cells ◽  
Phosphotransferase System ◽  
E Coli ◽  
Biochemical Effects ◽  
Inhibition Of Growth ◽  
Uncompetitive Inhibitor

The uptake of 4-deoxy-4-fluoro-D-glucose (4FG), without subsequent catabolism, by resting cells of Escherichia coli (ATCC 11775) is 0.06 mg/mg dry weight. In frozen–thawed cells of this organism, 4FG is a substrate for the phosphoenolpyruvate phosphotransferase system with a rate of phosphorylation twice that found for the isomeric 3-deoxy-3-fluoro-D-glucose. 4FG is not a carbon source for growth of this organism and it inhibits the extent of growth of cells in the presence of glucose. The inhibition of growth of E. coli K12 on lactose by 4FG is also observed and this is considered to be consistent with the fact that 4FG is an uncompetitive inhibitor of β-galactosidase (EC 3.2.1.23) activity and that 4FG or 4-deoxy-4-fluoro-D-glucose-6-phosphate repress β-galactosidase synthesis. These results support the view that catabolite repression may be produced by compounds which are not necessarily metabolised further than hexose-6-phosphates.

scholarly journals Constructing an ethanol utilization pathway in Escherichia coli to produce acetyl-CoA derived compounds

2020 ◽  
Author(s):  
Hong Liang ◽  
Xiaoqiang Ma ◽  
Wenbo Ning ◽  
Yurou Liu ◽  
Anthony J. Sinskey ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Carbon Source ◽  
Sole Carbon Source ◽  
Structural Similarity ◽  
List Type ◽  
Acetyl Coa ◽  
E Coli ◽  
Utilization Pathway ◽  
Aspartate Family

AbstractEngineering microbes to utilize non-conventional substrates could create short and efficient pathways to convert substrate into product. In this study, we designed and constructed a two-step heterologous ethanol utilization pathway (EUP) in Escherichia coli by using acetaldehyde dehydrogenase (encoded by ada) from Dickeya zeae and alcohol dehydrogenase (encoded by adh2) from Saccharomyces cerevisiae. This EUP can convert ethanol into acetyl-CoA without ATP consumption, and generate two molecules of NADH per molecule of ethanol. We optimized the expression of these two genes and found that ethanol consumption could be improved by expressing them in a specific order (ada-adh2) with a constitutive promoter (PgyrA). The engineered E. coli strain with EUP consumed approximately 8 g/L of ethanol in 96 hours when it was used as sole carbon source. Subsequently, we combined EUP with the biosynthesis of polyhydroxybutyrate (PHB), a biodegradable polymer derived from acetyl-CoA. The engineered E. coli strain carrying EUP and PHB biosynthetic pathway produced 1.1 g/L of PHB from 10 g/L of ethanol and 1 g/L of aspartate family amino acids in 96 hours. We also engineered E. coli strain to produced 24 mg/L of prenol from 10 g/L of ethanol in 48 hours, supporting the feasibility of converting ethanol into different classes of acetyl-CoA derived compounds.HighlightsEngineered Escherichia coli strains to grow on ethanol as sole carbon sourceDemonstrated that ethanol was converted into acetyl-CoA (AcCoA) through two pathways (acetaldehyde-acetate-AcCoA and acetaldehyde-AcCoA)Converted ethanol into two acetyl-CoA derived products with low structural similarity (polyhydroxybutyrate and prenol)Discovered that supplementation of the aspartate family amino acids can substantially improve cell growth on ethanol

scholarly journals Relaxation of catabolite repression in streptomycin-dependent Escherichia coli

1969 ◽  
Vol 111 (3) ◽  
pp. 279-286 ◽  
Author(s):  
M. B. Coukell ◽  
W. J. Polglase
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Catabolite Repression ◽  
Citrate Synthase ◽  
Wild Type ◽  
Experimental Conditions ◽  
E Coli ◽  
Equivalent Quantity ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Escherichia Coli B

Acetohydroxy acid synthetase, which is sensitive to catabolite repression in wild-type Escherichia coli B, was relatively resistant to this control in a streptomycin-dependent mutant. The streptomycin-dependent mutant was found to be inducible for β-galactosidase in the presence of glucose, although repression of β-galactosidase by glucose occurred under experimental conditions where growth of the streptomycin-dependent mutant was limited. Additional glucose-sensitive enzymes of wild-type E. coli B (citrate synthase, fumarase, aconitase and isocitrate dehydrogenase) were found to be insensitive to the carbon source in streptomycin-dependent mutants: these enzymes were formed by streptomycin-dependent E. coli B in equivalent quantities when either glucose or glycerol was the carbon source. Two enzymes, glucokinase and glucose 6-phosphate dehydrogenase, that are glucose-insensitive in wild-type E. coli B were formed in equivalent quantity on glucose or glycerol in both streptomycin-sensitive and streptomycin-dependent E. coli B. The results indicate a general decrease or relaxation of catabolite repression in the streptomycin-dependent mutant. The yield of streptomycin-dependent cells from glucose was one-third less than that of the streptomycin-sensitive strain. We conclude that the decreased efficiency of glucose utilization in streptomycin-dependent E. coli B is responsible for the relaxation of catabolite repression in this mutant.

scholarly journals DsdX Is the Second d-Serine Transporter in Uropathogenic Escherichia coli Clinical Isolate CFT073

2006 ◽  
Vol 188 (18) ◽  
pp. 6622-6628 ◽  
Author(s):  
Andrew T. Anfora ◽  
Rodney A. Welch
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Carbon Source ◽  
Clinical Isolate ◽  
Sole Carbon Source ◽  
Minimal Medium ◽  
Amino Acid Sequence Similarity ◽  
Predicted Amino Acid Sequence ◽  
E Coli ◽  
K 12

ABSTRACTd-Serine is an amino acid present in mammalian urine that is inhibitory toEscherichia colistrains lacking a functionaldsdAgene. Counterintuitively, adsdAstrain ofE. coliclinical isolate CFT073 hypercolonizes the bladder and kidneys of mice relative to wild type during a coinfection in the murine model of urinary tract infection. We are interested in the mechanisms for uptake ofd-serine in CFT073.d-Serine entersE. coliK-12 via CycA, thed-alanine transporter andd-cycloserine sensitivity locus. CFT073cycAcan grow on minimal medium withd-serine as a sole carbon source. ThedsdXgene of thedsdCXAlocus is a likely candidate for an additionald-serine transporter based on its predicted amino acid sequence similarity to gluconate transporters. In minimal medium, CFT073dsdXcan grow ond-serine as a sole carbon source; however, CFT073dsdX cycAcannot. Additionally, CFT073dsdXA cycAis not sensitive to inhibitory concentrations ofd-serine during growth on glycerol andd-serine minimal medium.d-[14C]serine uptake experiments with CFT073dsdX cycAharboringdsdXorcycArecombinant plasmids confirm thatd-serine is able to enterE. colicells via CycA or DsdX. In whole-celld-[14C]serine uptake experiments, DsdX has an apparentKmof 58.75 μM and aVmaxof 75.96 nmol/min/mg, and CycA has an apparentKmof 82.40 μM and aVmaxof 58.90 nmol/min/mg. Onlyd-threonine marginally inhibits DsdX-mediatedd-serine transport, whereasd-alanine, glycine, andd-cycloserine inhibit CycA-mediatedd-serine transport. DsdX or CycA is sufficient to transport physiological quantities ofd-serine, but DsdX is ad-serine-specific permease.

scholarly journals Identification, Source, and Metabolism of N-Ethylglutamate in Escherichia coli

2010 ◽  
Vol 192 (20) ◽  
pp. 5437-5440 ◽  
Author(s):  
Robert H. White
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Yeast Extract ◽  
Sole Carbon Source ◽  
Isotope Dilution Analysis ◽  
Sole Nitrogen Source ◽  
Glucose Medium ◽  
Detectable Amount ◽  
Deuterated Water ◽  
E Coli

ABSTRACT N-Ethylglutamate (NEG) was detected in Escherichia coli BL21 cells grown on LB broth, and it was found to occur at a concentration of ∼4 mM in these cells under these conditions. The same cells grown on M9 glucose medium contained no detectable amount of NEG. Analysis of the LB broth showed the presence of NEG, a compound never before reported as a natural product. Isotope dilution analysis showed that it occurred at a concentration of 160 μM in LB broth. Analyses of yeast extract and tryptone, the organic components of LB broth, both showed the presence NEG. It was demonstrated that NEG can be generated during the autolysis of the yeast used in the preparation of the yeast extract. Growth of these E. coli cells in LB broth prepared in deuterated water showed no incorporation of deuterium into NEG, demonstrating that E. coli cells did not generate the NEG. Cell growth rates were not affected by the addition of 5 mM NEG to either LB or M9 glucose medium. l-[ethyl-2H4]NEG was found to be readily incorporated into the cells and metabolized by the cells. From these results, it was concluded that all of the NEG present in the cells was taken up from the medium. NEG could serve as the sole nitrogen source for E. coli when grown on M9 glucose medium in the presence of glucose but could not serve as the sole carbon source on M9 medium in the absence of glucose.

scholarly journals The PalkBFGHJKL Promoter Is under Carbon Catabolite Repression Control in Pseudomonas oleovorans but Not in Escherichia coli alk+Recombinants

1999 ◽  
Vol 181 (5) ◽  
pp. 1610-1616 ◽  
Author(s):  
Ivo E. Staijen ◽  
Rosanna Marcionelli ◽  
Bernard Witholt
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Carbon Sources ◽  
Transcriptional Level ◽  
Pseudomonas Oleovorans ◽  
Hydroxylase Activity ◽  
Alkane Hydroxylase ◽  
Alkane Degradation ◽  
E Coli

ABSTRACT The alk genes are located on the OCT plasmid ofPseudomonas oleovorans and encode an inducible pathway for the utilization of n-alkanes as carbon and energy sources. We have investigated the influence of alternative carbon sources on the induction of this pathway in P. oleovorans andEscherichia coli alk + recombinants. In doing so, we confirmed earlier reports that induction of alkane hydroxylase activity in pseudomonads is subject to carbon catabolite repression. Specifically, synthesis of the monooxygenase component AlkB is repressed at the transcriptional level. The alk genes have been cloned into plasmid pGEc47, which has a copy number of about 5 to 10 per cell in both E. coli and pseudomonads.Pseudomonas putida GPo12 is a P. oleovoransderivative cured of the OCT plasmid. Upon introduction of pGEc47 in this strain, carbon catabolite repression of alkane hydroxylase activity was reduced significantly. In cultures of recombinant E. coli HB101 and W3110 carrying pGEc47, induction of AlkB and transcription of the alkB gene were no longer subject to carbon catabolite repression. This suggests that carbon catabolite repression of alkane degradation is regulated differently inPseudomonas and in E. coli strains. These results also indicate that P alkBFGHJKL , the P alk promoter, might be useful in attaining high expression levels of heterologous genes in E. coligrown on inexpensive carbon sources which normally trigger carbon catabolite repression of native expression systems in this host.

scholarly journals The Wide-Domain Carbon Catabolite Repressor CreA Indirectly Controls Expression of the Aspergillus nidulans xlnBGene, Encoding the Acidic Endo-β-(1,4)-Xylanase X24

2001 ◽  
Vol 183 (5) ◽  
pp. 1517-1523 ◽  
Author(s):  
Margarita Orejas ◽  
Andrew P. MacCabe ◽  
JoséAntonio Pérez-González ◽  
Sudeep Kumar ◽  
Daniel Ramón
Keyword(s):  
Carbon Source ◽  
Aspergillus Nidulans ◽  
Catabolite Repression ◽  
Sole Carbon Source ◽  
Carbon Catabolite Repression ◽  
Target Sites ◽  
Wide Domain ◽  
Catabolite Repressor ◽  
Functional Analyses

ABSTRACT The Aspergillus nidulans xlnB gene, which encodes the acidic endo-β-(1,4)-xylanase X24, is expressed when xylose is present as the sole carbon source and repressed in the presence of glucose. That the mutation creAd30results in considerably elevated levels of xlnB mRNA indicates a role for the wide-domain repressor CreA in the repression of xlnB promoter (xlnBp) activity. Functional analyses of xlnBp::goxC reporter constructs show that none of the four CreA consensus target sites identified in xlnBp are functional in vivo. The CreA repressor is thus likely to exert carbon catabolite repression via an indirect mechanism rather than to influence xlnB expression by acting directly on xlnB.

scholarly journals Metabolic engineering of Escherichia coli for de novo production of 3-phenylpropanol via retrobiosynthesis approach

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Zhenning Liu ◽  
Xue Zhang ◽  
Dengwei Lei ◽  
Bin Qiao ◽  
Guang-Rong Zhao
Keyword(s):  
Escherichia Coli ◽  
Metabolic Engineering ◽  
De Novo ◽  
Microbial Cell ◽  
Cell Factory ◽  
Phosphopantetheinyl Transferase ◽  
Chromosome Engineering ◽  
Microbial Cell Factory ◽  
E Coli ◽  
Artificial Pathway

Abstract Background 3-Phenylpropanol with a pleasant odor is widely used in foods, beverages and cosmetics as a fragrance ingredient. It also acts as the precursor and reactant in pharmaceutical and chemical industries. Currently, petroleum-based manufacturing processes of 3-phenypropanol is environmentally unfriendly and unsustainable. In this study, we aim to engineer Escherichia coli as microbial cell factory for de novo production of 3-phenypropanol via retrobiosynthesis approach. Results Aided by in silico retrobiosynthesis analysis, we designed a novel 3-phenylpropanol biosynthetic pathway extending from l-phenylalanine and comprising the phenylalanine ammonia lyase (PAL), enoate reductase (ER), aryl carboxylic acid reductase (CAR) and phosphopantetheinyl transferase (PPTase). We screened the enzymes from plants and microorganisms and reconstructed the artificial pathway for conversion of 3-phenylpropanol from l-phenylalanine. Then we conducted chromosome engineering to increase the supply of precursor l-phenylalanine and combined the upstream l-phenylalanine pathway and downstream 3-phenylpropanol pathway. Finally, we regulated the metabolic pathway strength and optimized fermentation conditions. As a consequence, metabolically engineered E. coli strain produced 847.97 mg/L of 3-phenypropanol at 24 h using glucose-glycerol mixture as co-carbon source. Conclusions We successfully developed an artificial 3-phenylpropanol pathway based on retrobiosynthesis approach, and highest titer of 3-phenylpropanol was achieved in E. coli via systems metabolic engineering strategies including enzyme sources variety, chromosome engineering, metabolic strength balancing and fermentation optimization. This work provides an engineered strain with industrial potential for production of 3-phenylpropanol, and the strategies applied here could be practical for bioengineers to design and reconstruct the microbial cell factory for high valuable chemicals.

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