c. glutamicum strains Search Results


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  • 99
    ATCC c glutamicum strains
    Representative anaerobic batch fermentation of C. <t>glutamicum</t> Δ ldhA /pAN6 -pyc P458S (♦; biomass concentration, 6.35 g cdw/liter), C. glutamicum BOL-1/pAN6 -pyc P458S (▲; biomass concentration, 6.00 g cdw/liter), and C. glutamicum BOL-2
    C Glutamicum Strains, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 200 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Difco c glutamicum strains
    Gas Chromatography/Mass Spectrometry (GC/MS) analysis of partially per- O -methylated, per- O -acetylated alditol acetate derivatives of purified arabinogalactan from C. <t>glutamicum</t> , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD . A , gas chromatograms demonstrating the presence of glycosyl linkages in purified AG. B , mass spectra fragmentation profile of peaks resolving at 11.3 min corresponding to 2-Ara f ( * ) and 3-Ara f ( ** ) and cleavage ions representing fragmentation of 2-Ara f ( * ) and 3-Ara f ( * ) located in terminal arabinan glycosyl motifs.
    C Glutamicum Strains, supplied by Difco, used in various techniques. Bioz Stars score: 94/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain f
    Transformation assays of FA and VAN. C. <t>glutamicum</t> <t>strain</t> F was cultured in BHI medium at 30 °C for 24 h, and 0.1 mL of each culture was transferred to 5 mL of AR medium containing 2 mM each of FA or VAN with 25 μg/mL of Km in 5 mL tubes. The biotransformation of a FA and b VAN was performed at 30 °C with agitation at 180 rpm for 72 h. The concentrations of extracellular FA ( squares ), VA ( triangles ), VAN ( diamonds ) and PCA ( circles ) were monitored every 24 h. Data represent the mean and standard error from three independent experiments
    C Glutamicum Strain F, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    TaKaRa c glutamicum strain r e coli strains hst02
    Transformation assays of FA and VAN. C. <t>glutamicum</t> <t>strain</t> F was cultured in BHI medium at 30 °C for 24 h, and 0.1 mL of each culture was transferred to 5 mL of AR medium containing 2 mM each of FA or VAN with 25 μg/mL of Km in 5 mL tubes. The biotransformation of a FA and b VAN was performed at 30 °C with agitation at 180 rpm for 72 h. The concentrations of extracellular FA ( squares ), VA ( triangles ), VAN ( diamonds ) and PCA ( circles ) were monitored every 24 h. Data represent the mean and standard error from three independent experiments
    C Glutamicum Strain R E Coli Strains Hst02, supplied by TaKaRa, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    ATCC c glutamicum strain dm 1698
    Transformation assays of FA and VAN. C. <t>glutamicum</t> <t>strain</t> F was cultured in BHI medium at 30 °C for 24 h, and 0.1 mL of each culture was transferred to 5 mL of AR medium containing 2 mM each of FA or VAN with 25 μg/mL of Km in 5 mL tubes. The biotransformation of a FA and b VAN was performed at 30 °C with agitation at 180 rpm for 72 h. The concentrations of extracellular FA ( squares ), VA ( triangles ), VAN ( diamonds ) and PCA ( circles ) were monitored every 24 h. Data represent the mean and standard error from three independent experiments
    C Glutamicum Strain Dm 1698, supplied by ATCC, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC strains c glutamicum atcc 21526
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), <t>ATCC</t> 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    Strains C Glutamicum Atcc 21526, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain atcc 14752
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), <t>ATCC</t> 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Strain Atcc 14752, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC strains c glutamicum atcc 13287
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), <t>ATCC</t> 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    Strains C Glutamicum Atcc 13287, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain atcc 21831
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), <t>ATCC</t> 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Strain Atcc 21831, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum corynebacterium glutamicum strain as019
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), <t>ATCC</t> 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Corynebacterium Glutamicum Strain As019, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain atcc 14067
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), <t>ATCC</t> 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Strain Atcc 14067, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain atcc 17965
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), <t>ATCC</t> 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Strain Atcc 17965, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum strain atcc 21850
    Fed-batch fermentation of violacein with C. <t>glutamicum</t> <t>ATCC</t> 21850 (pEC-C-vio1) in 3 L NBS BioFlo 115 fermenter
    C Glutamicum Strain Atcc 21850, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum atcc 31831 wild type strain
    galM and araR promoter activities. (A) araR promoter activity. The 400-bp araR upstream region was fused to the lacZ reporter gene in the canonical (P araR -F) or the reverse (P araR -R) orientation. The C. <t>glutamicum</t> <t>ATCC</t> 31831 strains carrying the respective
    C Glutamicum Atcc 31831 Wild Type Strain, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    ATCC wild type strain c glutamicum atcc 31833
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    Wild Type Strain C Glutamicum Atcc 31833, supplied by ATCC, used in various techniques. Bioz Stars score: 91/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    ATCC lysine producing c glutamicum atcc 21253 strains
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    Lysine Producing C Glutamicum Atcc 21253 Strains, supplied by ATCC, used in various techniques. Bioz Stars score: 91/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC l lysine producing strain c glutamicum atcc 21799
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    L Lysine Producing Strain C Glutamicum Atcc 21799, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Kyowa Hakko Kirin c glutamicum
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    C Glutamicum, supplied by Kyowa Hakko Kirin, used in various techniques. Bioz Stars score: 95/100, based on 48 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    ATCC c glutamicum atcc 13745
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    C Glutamicum Atcc 13745, supplied by ATCC, used in various techniques. Bioz Stars score: 91/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum atcc 13826
    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. <t>glutamicum</t> KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum <t>ATCC</t> 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.
    C Glutamicum Atcc 13826, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 32 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    ATCC c glutamicum atcc 21543
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and <t>ATCC</t> 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Atcc 21543, supplied by ATCC, used in various techniques. Bioz Stars score: 91/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC c glutamicum atcc 13803
    Impact of strain optimization of lysine-producing strains of C. <t>glutamicum</t> on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and <t>ATCC</t> 21543 (5). ICD, isocitrate dehydrogenase.
    C Glutamicum Atcc 13803, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Representative anaerobic batch fermentation of C. glutamicum Δ ldhA /pAN6 -pyc P458S (♦; biomass concentration, 6.35 g cdw/liter), C. glutamicum BOL-1/pAN6 -pyc P458S (▲; biomass concentration, 6.00 g cdw/liter), and C. glutamicum BOL-2

    Journal: Applied and Environmental Microbiology

    Article Title: Toward Homosuccinate Fermentation: Metabolic Engineering of Corynebacterium glutamicum for Anaerobic Production of Succinate from Glucose and Formate

    doi: 10.1128/AEM.07790-11

    Figure Lengend Snippet: Representative anaerobic batch fermentation of C. glutamicum Δ ldhA /pAN6 -pyc P458S (♦; biomass concentration, 6.35 g cdw/liter), C. glutamicum BOL-1/pAN6 -pyc P458S (▲; biomass concentration, 6.00 g cdw/liter), and C. glutamicum BOL-2

    Article Snippet: In this work, metabolic engineering was used to develop C. glutamicum strains (based on the ATCC 13032 wild-type strain) for anaerobic succinate production from glucose or glucose/formate mixtures.

    Techniques: Concentration Assay

    Schematic representation of the metabolic pathways relevant for succinate production by C. glutamicum under anaerobic conditions. Reactions leading to by-products or which are presumably not relevant are displayed in gray. Enzymes whose genes were deleted

    Journal: Applied and Environmental Microbiology

    Article Title: Toward Homosuccinate Fermentation: Metabolic Engineering of Corynebacterium glutamicum for Anaerobic Production of Succinate from Glucose and Formate

    doi: 10.1128/AEM.07790-11

    Figure Lengend Snippet: Schematic representation of the metabolic pathways relevant for succinate production by C. glutamicum under anaerobic conditions. Reactions leading to by-products or which are presumably not relevant are displayed in gray. Enzymes whose genes were deleted

    Article Snippet: In this work, metabolic engineering was used to develop C. glutamicum strains (based on the ATCC 13032 wild-type strain) for anaerobic succinate production from glucose or glucose/formate mixtures.

    Techniques:

    Representative anaerobic fed-batch fermentation with C. glutamicum strain BOL-3/pAN6 -gap showing succinate production (black stars) during coutilization of glucose (black squares) and formate (black triangles) (A) and formation of the by-products pyruvate

    Journal: Applied and Environmental Microbiology

    Article Title: Toward Homosuccinate Fermentation: Metabolic Engineering of Corynebacterium glutamicum for Anaerobic Production of Succinate from Glucose and Formate

    doi: 10.1128/AEM.07790-11

    Figure Lengend Snippet: Representative anaerobic fed-batch fermentation with C. glutamicum strain BOL-3/pAN6 -gap showing succinate production (black stars) during coutilization of glucose (black squares) and formate (black triangles) (A) and formation of the by-products pyruvate

    Article Snippet: In this work, metabolic engineering was used to develop C. glutamicum strains (based on the ATCC 13032 wild-type strain) for anaerobic succinate production from glucose or glucose/formate mixtures.

    Techniques:

    Minimizing acetate formation during anaerobic succinate production from glucose by C. glutamicum .

    Journal: Applied and Environmental Microbiology

    Article Title: Toward Homosuccinate Fermentation: Metabolic Engineering of Corynebacterium glutamicum for Anaerobic Production of Succinate from Glucose and Formate

    doi: 10.1128/AEM.07790-11

    Figure Lengend Snippet: Minimizing acetate formation during anaerobic succinate production from glucose by C. glutamicum .

    Article Snippet: In this work, metabolic engineering was used to develop C. glutamicum strains (based on the ATCC 13032 wild-type strain) for anaerobic succinate production from glucose or glucose/formate mixtures.

    Techniques:

    Assessment of different promoter activities in C. glutamicum ATCC 13032. (A) P BAD activities in C. glutamicum carrying pWYE1067- lacZ exposed to different l -arabinose concentrations. The l -arabinose concentrations are represented by different symbols:

    Journal: Applied and Environmental Microbiology

    Article Title: Development and Application of an Arabinose-Inducible Expression System by Facilitating Inducer Uptake in Corynebacterium glutamicum

    doi: 10.1128/AEM.01147-12

    Figure Lengend Snippet: Assessment of different promoter activities in C. glutamicum ATCC 13032. (A) P BAD activities in C. glutamicum carrying pWYE1067- lacZ exposed to different l -arabinose concentrations. The l -arabinose concentrations are represented by different symbols:

    Article Snippet: C. glutamicum strain ATCC 13032 was used for genetic disruption and expression using plasmid pK18 mobsacB and pXMJ19 derivatives ( , ).

    Techniques:

    Shake-flask fermentation profiles of C. glutamicum ATCC 13032 strain carrying the vector pWYE1088 (A) or the vector pWYE1088- odhI (B) under biotin nonlimiting conditions. l -Arabinose (0.02%) was used to induce odhI gene expression. The dry cell weight

    Journal: Applied and Environmental Microbiology

    Article Title: Development and Application of an Arabinose-Inducible Expression System by Facilitating Inducer Uptake in Corynebacterium glutamicum

    doi: 10.1128/AEM.01147-12

    Figure Lengend Snippet: Shake-flask fermentation profiles of C. glutamicum ATCC 13032 strain carrying the vector pWYE1088 (A) or the vector pWYE1088- odhI (B) under biotin nonlimiting conditions. l -Arabinose (0.02%) was used to induce odhI gene expression. The dry cell weight

    Article Snippet: C. glutamicum strain ATCC 13032 was used for genetic disruption and expression using plasmid pK18 mobsacB and pXMJ19 derivatives ( , ).

    Techniques: Plasmid Preparation, Expressing

    Characterization of the dynamic range of l -arabinose induction. (A) β-Galactosidase activities in C. glutamicum ATCC 13032 carrying pWYE1088- lacZ . Cells were harvested after 4 h of induction at the indicated l -arabinose concentrations for analysis.

    Journal: Applied and Environmental Microbiology

    Article Title: Development and Application of an Arabinose-Inducible Expression System by Facilitating Inducer Uptake in Corynebacterium glutamicum

    doi: 10.1128/AEM.01147-12

    Figure Lengend Snippet: Characterization of the dynamic range of l -arabinose induction. (A) β-Galactosidase activities in C. glutamicum ATCC 13032 carrying pWYE1088- lacZ . Cells were harvested after 4 h of induction at the indicated l -arabinose concentrations for analysis.

    Article Snippet: C. glutamicum strain ATCC 13032 was used for genetic disruption and expression using plasmid pK18 mobsacB and pXMJ19 derivatives ( , ).

    Techniques:

    Model of the pathway for assimilatory sulphur reduction in C. glutamicum . . For proteins with gene names given in black, the involvement in the reduction of sulphate has been verified experimentally, for those in grey it has been inferred from circumstantial evidence.

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: Model of the pathway for assimilatory sulphur reduction in C. glutamicum . . For proteins with gene names given in black, the involvement in the reduction of sulphate has been verified experimentally, for those in grey it has been inferred from circumstantial evidence.

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques:

    Conservation of the C. glutamicum gene cluster possibly involved in assimilatory sulphate reduction in the Actinomycetales . Functions were inferred based on sequence similarity from BLAST searches against the UniProt database. Only those genes are displayed that were found to be clustered (at least two adjacent genes possibly involved in assimilatory sulphate reduction).

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: Conservation of the C. glutamicum gene cluster possibly involved in assimilatory sulphate reduction in the Actinomycetales . Functions were inferred based on sequence similarity from BLAST searches against the UniProt database. Only those genes are displayed that were found to be clustered (at least two adjacent genes possibly involved in assimilatory sulphate reduction).

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques: Sequencing

    Growth of the C. glutamicum wild-type and the ΔcysZ mutant strain in medium with different sulphate concentrations . Wild-type and mutant strain were grown in liquid minimal medium, cell growth was determined by real-time nephelometry. For each time point the mean of 18 measurements is displayed (3 independent cultivations, 6 parallels per cultivation). The wild-type strain is denoted with filled diamonds, open symbols are used to indicate the mutant strain.

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: Growth of the C. glutamicum wild-type and the ΔcysZ mutant strain in medium with different sulphate concentrations . Wild-type and mutant strain were grown in liquid minimal medium, cell growth was determined by real-time nephelometry. For each time point the mean of 18 measurements is displayed (3 independent cultivations, 6 parallels per cultivation). The wild-type strain is denoted with filled diamonds, open symbols are used to indicate the mutant strain.

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques: Mutagenesis

    Comparison of the mRNA levels of the cys genes of the C. glutamicum wild-type and a cysI transposon mutant . Total RNA was isolated from cells grown in MMS with 1 mM L-cysteine as sulphur source and the relative transcription levels were determined using real-time RT-PCR to quantify the mRNAs of the displayed genes. Small black bars inside the arrows representing the genes indicate the position of the internal fragments amplified in the real-time RT-PCR.

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: Comparison of the mRNA levels of the cys genes of the C. glutamicum wild-type and a cysI transposon mutant . Total RNA was isolated from cells grown in MMS with 1 mM L-cysteine as sulphur source and the relative transcription levels were determined using real-time RT-PCR to quantify the mRNAs of the displayed genes. Small black bars inside the arrows representing the genes indicate the position of the internal fragments amplified in the real-time RT-PCR.

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques: Mutagenesis, Isolation, Quantitative RT-PCR, Amplification

    The promoter/operator regions of the C. glutamicum fpr2 and cysI genes . The determined transcriptional start points of the two transcriptional units fpr2 (A) and cysIXHDNYZ (B) are marked as '+1'. Parts of the two potential promoters (-35, -10, +1) are overlined, bases in bold type in these regions indicate bases matching the C. glutamicum σ 70 consensus promoter [23]. DNA motifs matching the consensus sequence of the McbR binding-site are boxed. Bases in bold italics mark potential ribosome binding-sites, open underlining arrows indicate the annotated starts of genes.

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: The promoter/operator regions of the C. glutamicum fpr2 and cysI genes . The determined transcriptional start points of the two transcriptional units fpr2 (A) and cysIXHDNYZ (B) are marked as '+1'. Parts of the two potential promoters (-35, -10, +1) are overlined, bases in bold type in these regions indicate bases matching the C. glutamicum σ 70 consensus promoter [23]. DNA motifs matching the consensus sequence of the McbR binding-site are boxed. Bases in bold italics mark potential ribosome binding-sites, open underlining arrows indicate the annotated starts of genes.

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques: Sequencing, Binding Assay

    The fpr2cysIXHDNYZ gene cluster in the C. glutamicum genome . Coloured arrows indicate genes that are part of the cluster most probably involved in assimilatory sulphate reduction. Hairpins mark potential rho-independent transcription termination signals predicted by the TransTerm software. Black bars denote binding sites for the transcriptional repressor McbR [15].

    Journal: BMC Genomics

    Article Title: Functional genomics and expression analysis of the Corynebacterium glutamicum fpr2-cysIXHDNYZ gene cluster involved in assimilatory sulphate reduction

    doi: 10.1186/1471-2164-6-121

    Figure Lengend Snippet: The fpr2cysIXHDNYZ gene cluster in the C. glutamicum genome . Coloured arrows indicate genes that are part of the cluster most probably involved in assimilatory sulphate reduction. Hairpins mark potential rho-independent transcription termination signals predicted by the TransTerm software. Black bars denote binding sites for the transcriptional repressor McbR [15].

    Article Snippet: The plasmids pCR018d to pCR025d were transferred into the C. glutamicum wild-type strain (ATCC 13032) by electroporation [ ].

    Techniques: Software, Binding Assay

    The growth (A) , glucose consumption (B) and putrescine production (C) in Corynebacterium glutamicum PUT-ALE (•) and ATCC 13032 (∘). Data represent the average of three replicates and error bars represent standard deviation.

    Journal: Frontiers in Microbiology

    Article Title: Transcriptomic Changes in Response to Putrescine Production in Metabolically Engineered Corynebacterium glutamicum

    doi: 10.3389/fmicb.2017.01987

    Figure Lengend Snippet: The growth (A) , glucose consumption (B) and putrescine production (C) in Corynebacterium glutamicum PUT-ALE (•) and ATCC 13032 (∘). Data represent the average of three replicates and error bars represent standard deviation.

    Article Snippet: We first compared the growth, glucose consumption, and putrescine production of the C. glutamicum PUT-ALE and the wild-type strain C. glutamicum ATCC 13032 strains.

    Techniques: Standard Deviation

    Differentially expressed genes involved in glycolysis, the TCA cycle, pyruvate metabolism, amino acid biosynthesis and the putrescine biosynthetic pathway. The numbers indicate the values of the log 2 ratios of the expression levels in C. glutamicum PUT-ALE vs. C. glutamicum ATCC 13032. Red indicates upregulation. Blue indicates downregulation. Glc, glucose; G6P, glucose 6-phosphate; F6P, fructose 6-phosphate; F1,6P, fructose 1,6-bisphosphate; GAP, D -Glyceraldehyde 3-phosphate; GlyP, glycerone phosphate; G1,3P, 1,3-bisphospho- D -glycerate; G3P, 3-phosphoglycerate; G2P, 2-phospho-(R)-glycerate; PEP, phosphoenolpyruvate; Pyr, pyruvate; AcCoA, acetyl-CoA; GlcLac, D -glucono-1,5-lactone 6-phosphate; 6- P -glucon, 6-phospho- D -gluconate; Ribu5P, D -Ribulose 5-phosphate; Rib5P, D -ribose 5-phosphate; Xyl5P, D -Xylulose 5-phosphate; S7P, D -sedoheptulose 7-phosphate; E4P, D -erythrose 4-phosphate; PRPP, 5-phosphoribosyl diphosphate; His, L -histidine; DAHP, 3-deoxy-arabino-heptulonate 7-phosphate; Trp, L -tryptophan; Phe, L -phenylalanine; Tyr: L -tyrosine; D -Lac, D -Lactate; L -Lac, L -lactate; Ace, acetate; Val, L -valine; Ile, L -isoleucine; Leu, L -leucine; Ser, L -serine; Gly, L -glycine; Cys, L -cysteine; Ala, L -alanine; Cit, citrate; Ici, isocitrate; KG, 2-oxoglutarate; SucCoA, succinyl-CoA; Suc, succinate; Fum, fumarate; Mal, malate; OAA, oxaloacetate; Asp, L -aspartate; Asn, L -asparagine; ASA, L-aspartate 4-semialdehyde; HTPA, (2S,4S)-4-hydroxy-2,3,4,5-tetrahydrodipicolinate; Lys, L -lysine; Hom, homoserine; Thr, L -threonine; Ac-Hom, O -acetylhomoserine; Homcys, L -homocysteine; Met, L -methionine. Glut, L -glutamate; Gln, L -glutamine; GlutP, L -glutamate 5-phosphate; NAGlut, N -acetylglutamate; NAGlutP, N -acetyl-glutamyl 5-phosphate; NAGlut-semialdehyde, N -acetylglutamate semialdehyde; NAOrn, N -acetyl-ornithine; Orn, ornithine; Arg, L -arginine.

    Journal: Frontiers in Microbiology

    Article Title: Transcriptomic Changes in Response to Putrescine Production in Metabolically Engineered Corynebacterium glutamicum

    doi: 10.3389/fmicb.2017.01987

    Figure Lengend Snippet: Differentially expressed genes involved in glycolysis, the TCA cycle, pyruvate metabolism, amino acid biosynthesis and the putrescine biosynthetic pathway. The numbers indicate the values of the log 2 ratios of the expression levels in C. glutamicum PUT-ALE vs. C. glutamicum ATCC 13032. Red indicates upregulation. Blue indicates downregulation. Glc, glucose; G6P, glucose 6-phosphate; F6P, fructose 6-phosphate; F1,6P, fructose 1,6-bisphosphate; GAP, D -Glyceraldehyde 3-phosphate; GlyP, glycerone phosphate; G1,3P, 1,3-bisphospho- D -glycerate; G3P, 3-phosphoglycerate; G2P, 2-phospho-(R)-glycerate; PEP, phosphoenolpyruvate; Pyr, pyruvate; AcCoA, acetyl-CoA; GlcLac, D -glucono-1,5-lactone 6-phosphate; 6- P -glucon, 6-phospho- D -gluconate; Ribu5P, D -Ribulose 5-phosphate; Rib5P, D -ribose 5-phosphate; Xyl5P, D -Xylulose 5-phosphate; S7P, D -sedoheptulose 7-phosphate; E4P, D -erythrose 4-phosphate; PRPP, 5-phosphoribosyl diphosphate; His, L -histidine; DAHP, 3-deoxy-arabino-heptulonate 7-phosphate; Trp, L -tryptophan; Phe, L -phenylalanine; Tyr: L -tyrosine; D -Lac, D -Lactate; L -Lac, L -lactate; Ace, acetate; Val, L -valine; Ile, L -isoleucine; Leu, L -leucine; Ser, L -serine; Gly, L -glycine; Cys, L -cysteine; Ala, L -alanine; Cit, citrate; Ici, isocitrate; KG, 2-oxoglutarate; SucCoA, succinyl-CoA; Suc, succinate; Fum, fumarate; Mal, malate; OAA, oxaloacetate; Asp, L -aspartate; Asn, L -asparagine; ASA, L-aspartate 4-semialdehyde; HTPA, (2S,4S)-4-hydroxy-2,3,4,5-tetrahydrodipicolinate; Lys, L -lysine; Hom, homoserine; Thr, L -threonine; Ac-Hom, O -acetylhomoserine; Homcys, L -homocysteine; Met, L -methionine. Glut, L -glutamate; Gln, L -glutamine; GlutP, L -glutamate 5-phosphate; NAGlut, N -acetylglutamate; NAGlutP, N -acetyl-glutamyl 5-phosphate; NAGlut-semialdehyde, N -acetylglutamate semialdehyde; NAOrn, N -acetyl-ornithine; Orn, ornithine; Arg, L -arginine.

    Article Snippet: We first compared the growth, glucose consumption, and putrescine production of the C. glutamicum PUT-ALE and the wild-type strain C. glutamicum ATCC 13032 strains.

    Techniques: Expressing, Gas Chromatography

    CgpS is a prophage-encoded nucleoid-associated protein in C. glutamicum ATCC 13032. ( A ) Genomic organization of the CGP3 prophage region containing the cgpS gene in C. glutamicum strain ATCC 13032. ( B ) DNA affinity chromatography was used to identify putative regulatory proteins bound to the early phage promoter of alpAC ( 34 ). Protein samples were separated by SDS-PAGE, and enriched proteins were identified using MALD-ToF analysis. The nucleoid-associated protein CgpS (Cg1966) was detected as a prominent band with a molecular mass of 13.4 kDa. ( C ) Amino acid sequences of CgpS, five related proteins of Corynebacteria and five Lsr2 homologs of Mycobacteria were used for a multiple sequence alignment conducted using the Clustal Omega platform ( 73 ). The predicted domain organization is depicted above the sequence alignment. The Blosum62 identity score is highlighted in blue and matches between 21–26% across the species. The secondary structure of CgpS was predicted with 99.7% confidence of 54 residues (46% of CgpS sequence) by Phyre 2 ( 74 ). The motif ‘RGI,’ which is similar to the AT hook motif ‘RGR’ of Lsr2 and H-NS ( 44 , 59 ), was identified between two predicted alpha helices.

    Journal: Nucleic Acids Research

    Article Title: Silencing of cryptic prophages in Corynebacterium glutamicum

    doi: 10.1093/nar/gkw692

    Figure Lengend Snippet: CgpS is a prophage-encoded nucleoid-associated protein in C. glutamicum ATCC 13032. ( A ) Genomic organization of the CGP3 prophage region containing the cgpS gene in C. glutamicum strain ATCC 13032. ( B ) DNA affinity chromatography was used to identify putative regulatory proteins bound to the early phage promoter of alpAC ( 34 ). Protein samples were separated by SDS-PAGE, and enriched proteins were identified using MALD-ToF analysis. The nucleoid-associated protein CgpS (Cg1966) was detected as a prominent band with a molecular mass of 13.4 kDa. ( C ) Amino acid sequences of CgpS, five related proteins of Corynebacteria and five Lsr2 homologs of Mycobacteria were used for a multiple sequence alignment conducted using the Clustal Omega platform ( 73 ). The predicted domain organization is depicted above the sequence alignment. The Blosum62 identity score is highlighted in blue and matches between 21–26% across the species. The secondary structure of CgpS was predicted with 99.7% confidence of 54 residues (46% of CgpS sequence) by Phyre 2 ( 74 ). The motif ‘RGI,’ which is similar to the AT hook motif ‘RGR’ of Lsr2 and H-NS ( 44 , 59 ), was identified between two predicted alpha helices.

    Article Snippet: To test the proposed exclusion mechanism, we expressed the hns gene from E. coli MG1655 in a C. glutamicum ATCC 13032 strain containing the prophage reporter (::Plys -eyfp ).

    Techniques: Affinity Chromatography, SDS Page, Sequencing

    Countersilencing by overexpression of the N-terminal oligomerization domain of CgpS. ( A ) Growth studies of the wild-type ATCC 13032 and a strain lacking the CGP3 prophage (ΔCGP3), both carrying the pAN6-N- cgpS overexpression plasmid. Strains were grown in CGXII minimal medium with and without IPTG (150 μM). Data represent average values and standard deviations of three biological replicates. ( B ) Relative quantification of CGP3 excision using qPCR ( 20 ). The N-terminal domain of CgpS and the full-length protein were overproduced as described in (A). Samples for qPCR analysis were taken after 24 h. The relative amounts of circular phage DNA of induced and uninduced samples were compared. As a control wild-type cells with the empty plasmid were used. Data represent average values and standard deviations of three biological replicates. ( C ) Time-lapse fluorescence microscopy of the C. glutamicum prophage reporter strain (ATCC 13032::P lys-eyfp ) carrying the pAN6- N-cgpS . Cells were grown in PDMS-based microfluidic chip devices under continuous supply of CGXII with 25 μg·ml −1 kanamycin and with or without 150 μM IPTG to induce the expression of the truncated CpgS variant ( 36 ) (300 nl·min −1 ) (Video S1 and S2). ( D ) Comparative transcriptome analysis of C. glutamicum ATCC 13032 containing the overexpression plasmid pAN6- N-cgpS and a strain containing the empty vector control was performed as described in the Materials and Methods section.

    Journal: Nucleic Acids Research

    Article Title: Silencing of cryptic prophages in Corynebacterium glutamicum

    doi: 10.1093/nar/gkw692

    Figure Lengend Snippet: Countersilencing by overexpression of the N-terminal oligomerization domain of CgpS. ( A ) Growth studies of the wild-type ATCC 13032 and a strain lacking the CGP3 prophage (ΔCGP3), both carrying the pAN6-N- cgpS overexpression plasmid. Strains were grown in CGXII minimal medium with and without IPTG (150 μM). Data represent average values and standard deviations of three biological replicates. ( B ) Relative quantification of CGP3 excision using qPCR ( 20 ). The N-terminal domain of CgpS and the full-length protein were overproduced as described in (A). Samples for qPCR analysis were taken after 24 h. The relative amounts of circular phage DNA of induced and uninduced samples were compared. As a control wild-type cells with the empty plasmid were used. Data represent average values and standard deviations of three biological replicates. ( C ) Time-lapse fluorescence microscopy of the C. glutamicum prophage reporter strain (ATCC 13032::P lys-eyfp ) carrying the pAN6- N-cgpS . Cells were grown in PDMS-based microfluidic chip devices under continuous supply of CGXII with 25 μg·ml −1 kanamycin and with or without 150 μM IPTG to induce the expression of the truncated CpgS variant ( 36 ) (300 nl·min −1 ) (Video S1 and S2). ( D ) Comparative transcriptome analysis of C. glutamicum ATCC 13032 containing the overexpression plasmid pAN6- N-cgpS and a strain containing the empty vector control was performed as described in the Materials and Methods section.

    Article Snippet: To test the proposed exclusion mechanism, we expressed the hns gene from E. coli MG1655 in a C. glutamicum ATCC 13032 strain containing the prophage reporter (::Plys -eyfp ).

    Techniques: Over Expression, Plasmid Preparation, Real-time Polymerase Chain Reaction, Fluorescence, Microscopy, Chromatin Immunoprecipitation, Expressing, Variant Assay

    CgpS functions as a silencer of CGP3 prophage activity. ( A ) Silencing of CGP3 induction. The prophage reporter strain C. glutamicum ATCC 13032::P lys -eyfp containing the cgpS overexpression plasmid pAN6- cgpS-Strep was cultivated in CGXII minimal medium in the presence or absence of IPTG (50 μM) and MMC (600 μM). The prophage reporter strain carrying the empty plasmid pAN6 served as a control. EYFP fluorescence was measured after 20 h of microplate cultivation. ( B and C ) CgpS is located in the nucleoid and displays colocalization with the phage adaptor protein AlpA ( 34 ). For co-localization studies, a C-terminal CgpS-mCherry fusion and a C-terminal fusion of the prophage adaptor protein AlpA to eYFP (pEC-XC99E) were analyzed. Both gene fusions were under control of the inducible tac promoter. DNA was stained with Hoechst Dye 33342. Fluorescence images were taken after 2.5 h of cultivation in CGXII with 150 μM IPTG. Scale bar, 2 μm.

    Journal: Nucleic Acids Research

    Article Title: Silencing of cryptic prophages in Corynebacterium glutamicum

    doi: 10.1093/nar/gkw692

    Figure Lengend Snippet: CgpS functions as a silencer of CGP3 prophage activity. ( A ) Silencing of CGP3 induction. The prophage reporter strain C. glutamicum ATCC 13032::P lys -eyfp containing the cgpS overexpression plasmid pAN6- cgpS-Strep was cultivated in CGXII minimal medium in the presence or absence of IPTG (50 μM) and MMC (600 μM). The prophage reporter strain carrying the empty plasmid pAN6 served as a control. EYFP fluorescence was measured after 20 h of microplate cultivation. ( B and C ) CgpS is located in the nucleoid and displays colocalization with the phage adaptor protein AlpA ( 34 ). For co-localization studies, a C-terminal CgpS-mCherry fusion and a C-terminal fusion of the prophage adaptor protein AlpA to eYFP (pEC-XC99E) were analyzed. Both gene fusions were under control of the inducible tac promoter. DNA was stained with Hoechst Dye 33342. Fluorescence images were taken after 2.5 h of cultivation in CGXII with 150 μM IPTG. Scale bar, 2 μm.

    Article Snippet: To test the proposed exclusion mechanism, we expressed the hns gene from E. coli MG1655 in a C. glutamicum ATCC 13032 strain containing the prophage reporter (::Plys -eyfp ).

    Techniques: Activity Assay, Over Expression, Plasmid Preparation, Fluorescence, Staining

    Live/dead staining of C. glutamicum cells using the fluorescent dyes PI and Syto 9. C. glutamicum ATCC 13032 was grown on CGXII minimal medium with 4% glucose, sampled as indicated and stained with PI (red) and Syto 9 (green) as described in Experimental procedures .A. Calibration: The assumed fraction of dead (isopropanol-killed) cells was plotted against the measured fraction of dead cells.B. Percentage of Syto 9 and Syto 9/PI positive (live) and PI positive (dead) cells in dependency of cultivation time. Bars represent the mean ± SD of three independent experiments.C. Dot plots (PI versus Syto 9) of the control experiment (calibration), early log, log, and stationary phase C. glutamicum cells (live: green; dead: red).D. Microscopic analysis (fluorescence channel) of log phase C. glutamicum cells (5.5 h after inoculation) stained with Syto 9 and PI. The majority of cells showed a green fluorescence (I), a small amount of cells stained PI positive (II). Often V-shaped sister cells were observed where one cell stained Syto 9 positive whereas the sister cell stained PI positive (III/IV).

    Journal: Microbial Biotechnology

    Article Title: Monitoring of population dynamics of Corynebacterium glutamicum by multiparameter flow cytometry

    doi: 10.1111/1751-7915.12018

    Figure Lengend Snippet: Live/dead staining of C. glutamicum cells using the fluorescent dyes PI and Syto 9. C. glutamicum ATCC 13032 was grown on CGXII minimal medium with 4% glucose, sampled as indicated and stained with PI (red) and Syto 9 (green) as described in Experimental procedures .A. Calibration: The assumed fraction of dead (isopropanol-killed) cells was plotted against the measured fraction of dead cells.B. Percentage of Syto 9 and Syto 9/PI positive (live) and PI positive (dead) cells in dependency of cultivation time. Bars represent the mean ± SD of three independent experiments.C. Dot plots (PI versus Syto 9) of the control experiment (calibration), early log, log, and stationary phase C. glutamicum cells (live: green; dead: red).D. Microscopic analysis (fluorescence channel) of log phase C. glutamicum cells (5.5 h after inoculation) stained with Syto 9 and PI. The majority of cells showed a green fluorescence (I), a small amount of cells stained PI positive (II). Often V-shaped sister cells were observed where one cell stained Syto 9 positive whereas the sister cell stained PI positive (III/IV).

    Article Snippet: The strain C. glutamicum ATCC 13032 is known to contain one circular chromosome (Kalinowski et al ., ).

    Techniques: Staining, Fluorescence

    Growth of C. glutamicum ATCC 13032 on (A) CGXII minimal medium with 4% glucose and (B) BHI complex medium in a 50 ml shaking flask culture at 30°C and 120 r.p.m.

    Journal: Microbial Biotechnology

    Article Title: Monitoring of population dynamics of Corynebacterium glutamicum by multiparameter flow cytometry

    doi: 10.1111/1751-7915.12018

    Figure Lengend Snippet: Growth of C. glutamicum ATCC 13032 on (A) CGXII minimal medium with 4% glucose and (B) BHI complex medium in a 50 ml shaking flask culture at 30°C and 120 r.p.m.

    Article Snippet: The strain C. glutamicum ATCC 13032 is known to contain one circular chromosome (Kalinowski et al ., ).

    Techniques:

    Colocalization of Venus-LcpA and EYFP-LcpB with newly synthesized peptidoglycan in C. glutamicum ATCC 13032. The cells carrying a plasmid encoding Venus N-terminally fused to LcpA (pAN6-venus- lcpA ) or EYFP N-terminally fused to LcpB (pAN6-eyfp- lcpB ) were

    Journal: Journal of Bacteriology

    Article Title: Impact of LytR-CpsA-Psr Proteins on Cell Wall Biosynthesis in Corynebacterium glutamicum

    doi: 10.1128/JB.00406-16

    Figure Lengend Snippet: Colocalization of Venus-LcpA and EYFP-LcpB with newly synthesized peptidoglycan in C. glutamicum ATCC 13032. The cells carrying a plasmid encoding Venus N-terminally fused to LcpA (pAN6-venus- lcpA ) or EYFP N-terminally fused to LcpB (pAN6-eyfp- lcpB ) were

    Article Snippet: The C. glutamicum type strain ATCC 13032 was used as the wild type.

    Techniques: Synthesized, Plasmid Preparation

    Overview of the transcriptional and translational regulation in L-valine production strain C. glutamicum VWB-1. Red box repesents the gene was up-regulated in transcriptional level; Green box represents the gene was down-regulated in transcriptional level; blue box represents the gene was up-regulated both in transcriptional and translational level; rose red box represents the gene was down-regulated both in transcriptional and translational level. Figure 6 was drawn by Hailing Zhang, the first author of this article.

    Journal: Scientific Reports

    Article Title: Understanding the high l-valine production in Corynebacterium glutamicum VWB-1 using transcriptomics and proteomics

    doi: 10.1038/s41598-018-21926-5

    Figure Lengend Snippet: Overview of the transcriptional and translational regulation in L-valine production strain C. glutamicum VWB-1. Red box repesents the gene was up-regulated in transcriptional level; Green box represents the gene was down-regulated in transcriptional level; blue box represents the gene was up-regulated both in transcriptional and translational level; rose red box represents the gene was down-regulated both in transcriptional and translational level. Figure 6 was drawn by Hailing Zhang, the first author of this article.

    Article Snippet: Hence, the whole genome sequencing and annotation of the type strain C. glutamicum ATCC 13032 and the wild-type strain C. glutamicum R have deeply influenced the bacterial fermentation performance promotion during the last few years – .

    Techniques:

    Experimental design for quantification of flux parameters at the pyruvate node of Corynebacterium glutamicum with an equimolar mixture of [ 13 C 6 ] glucose and naturally labelled glucose . Relative change of the mass isotopomer distribution of aspartate (m/z 418) with varied Φ PEPC (A), relative change of the mass isotopomer distribution of valine (m/z 288) with varied ζ PEPC/PEPCK (B), relative change of the mass isotopomer distribution of alanine (m/z 260) with varied ζ PC/MAE (C). The labelling patterns at sole contribution of PEPC (Φ PEPC = 100 %), and highly reversible fluxes at the pyruvate node (ζ PEPC/PEPCK = ζ PC/MAE = 10) are taken as reference point and set to 100 %. The flux parameters investigated here comprise Φ PEPC (flux partitioning between PEPC and PC), ζ PEPC/PEPCK (ratio of exchange flux to net flux between the pools of PEP and OAA/MAL catalyzed by PEPC and PEPCK) and ζ PC/MAE (ratio of exchange flux to net flux between the pools of PYR and OAA/MAL catalyzed by PC and MAE). The exact definitions for the flux parameters are given in the appendix. Unless varied the flux parameters reflect the situation for the parent strain C. glutamicum lysC fbr (Figure 4).

    Journal: Microbial Cell Factories

    Article Title: Metabolic responses to pyruvate kinase deletion in lysine producing Corynebacterium glutamicum

    doi: 10.1186/1475-2859-7-8

    Figure Lengend Snippet: Experimental design for quantification of flux parameters at the pyruvate node of Corynebacterium glutamicum with an equimolar mixture of [ 13 C 6 ] glucose and naturally labelled glucose . Relative change of the mass isotopomer distribution of aspartate (m/z 418) with varied Φ PEPC (A), relative change of the mass isotopomer distribution of valine (m/z 288) with varied ζ PEPC/PEPCK (B), relative change of the mass isotopomer distribution of alanine (m/z 260) with varied ζ PC/MAE (C). The labelling patterns at sole contribution of PEPC (Φ PEPC = 100 %), and highly reversible fluxes at the pyruvate node (ζ PEPC/PEPCK = ζ PC/MAE = 10) are taken as reference point and set to 100 %. The flux parameters investigated here comprise Φ PEPC (flux partitioning between PEPC and PC), ζ PEPC/PEPCK (ratio of exchange flux to net flux between the pools of PEP and OAA/MAL catalyzed by PEPC and PEPCK) and ζ PC/MAE (ratio of exchange flux to net flux between the pools of PYR and OAA/MAL catalyzed by PC and MAE). The exact definitions for the flux parameters are given in the appendix. Unless varied the flux parameters reflect the situation for the parent strain C. glutamicum lysC fbr (Figure 4).

    Article Snippet: Microorganisms The reference strain C. glutamicum ATCC 13032 was derived from the American Type and Culture Collection (Manassas, USA).

    Techniques:

    Southern blot analysis of C. glutamicum wild-type ATCC 13032 and C. glutamicum ATCC 13032Δ porA cells. (A) Chromosomal DNA from wild-type cells (wt) and the Δ porA mutant strain (Δ porA ) were digested with restriction enzyme Bgl I, subjected to agarose gel electrophoresis, transferred onto a nylon membrane, and probed with a digoxigenin-labeled oligonucleotide corresponding to the last 33 bp of the porA gene. (B) Schematic representation from the 169-bp fragment released in the chromosome of the wild-type C. glutamicum strain after cleavage with Bgl I. Dig, digoxigenin

    Journal: Journal of Bacteriology

    Article Title: PorA Represents the Major Cell Wall Channel of the Gram-Positive Bacterium Corynebacterium glutamicum

    doi: 10.1128/JB.185.16.4779-4786.2003

    Figure Lengend Snippet: Southern blot analysis of C. glutamicum wild-type ATCC 13032 and C. glutamicum ATCC 13032Δ porA cells. (A) Chromosomal DNA from wild-type cells (wt) and the Δ porA mutant strain (Δ porA ) were digested with restriction enzyme Bgl I, subjected to agarose gel electrophoresis, transferred onto a nylon membrane, and probed with a digoxigenin-labeled oligonucleotide corresponding to the last 33 bp of the porA gene. (B) Schematic representation from the 169-bp fragment released in the chromosome of the wild-type C. glutamicum strain after cleavage with Bgl I. Dig, digoxigenin

    Article Snippet: The channel-forming polypeptide PorA is a 45-amino-acid acidic polypeptide with an excess of four negatively charged amino acids, which is encoded by the 138-bp gene porA . porA was deleted from the chromosome of C.glutamicum wild-type strain ATCC 13032 to obtain mutant ATCC 13032Δ porA .

    Techniques: Southern Blot, Mutagenesis, Agarose Gel Electrophoresis, Labeling

    Biosensor-based online monitoring of L-valine production in PDHC-deficient C. glutamicum strains. ( A ) Growth and ( B ) Lrp-sensor output (eYFP fluorescence) of the sensor strains C. glutamicum ATCC 13032 wild type (stars), ΔaceE (diamonds), ΔaceE Δpqo (circles), ΔaceE Δpqo Δpgi (triangles), and ΔaceE Δpqo Δpgi Δpyc (squares) cultivated in CGXII minimal medium containing 222 mM glucose and 154 mM acetate. Data represent average values of three independent cultivations. The transition of the producer strains into the stationary and production phase is highlighted by the grey area. ( C ) EYFP fluorescence of respective strains at the beginning of the production phase (black bars) and twelve hours after the initiation of L-valine production (grey bars). L-valine concentration (mM) in the supernatant of the respective strain 25 h after beginning of cultivation as measured by HPLC is indicated above the grey bars.

    Journal: PLoS ONE

    Article Title: Application of a Genetically Encoded Biosensor for Live Cell Imaging of L-Valine Production in Pyruvate Dehydrogenase Complex-Deficient Corynebacterium glutamicum Strains

    doi: 10.1371/journal.pone.0085731

    Figure Lengend Snippet: Biosensor-based online monitoring of L-valine production in PDHC-deficient C. glutamicum strains. ( A ) Growth and ( B ) Lrp-sensor output (eYFP fluorescence) of the sensor strains C. glutamicum ATCC 13032 wild type (stars), ΔaceE (diamonds), ΔaceE Δpqo (circles), ΔaceE Δpqo Δpgi (triangles), and ΔaceE Δpqo Δpgi Δpyc (squares) cultivated in CGXII minimal medium containing 222 mM glucose and 154 mM acetate. Data represent average values of three independent cultivations. The transition of the producer strains into the stationary and production phase is highlighted by the grey area. ( C ) EYFP fluorescence of respective strains at the beginning of the production phase (black bars) and twelve hours after the initiation of L-valine production (grey bars). L-valine concentration (mM) in the supernatant of the respective strain 25 h after beginning of cultivation as measured by HPLC is indicated above the grey bars.

    Article Snippet: Growth and production of C. glutamicum ATCC 13032 ΔaceE sensor strain.

    Techniques: Fluorescence, Concentration Assay, High Performance Liquid Chromatography

    Adaptive evolution experiment. ( A ) C. glutamicum ATCC 13032 and its prophage-free variant MB001 were adaptively evolved towards increased growth rates on glucose minimal medium (CGXII medium with 2% (w/v) glucose). Overall, six independent cell lines for each strain were evolved by repetitive batch cultivation using 48-well plates incubated at 30 °C (experimental details are given in the material and methods section). ( B ) Time line of the ALE experiment. Highlighted are the samples that were further analyzed by genome sequencing. Samples of the second reproduction of the experiment are marked with an asterisk.

    Journal: Scientific Reports

    Article Title: Adaptive laboratory evolution of Corynebacterium glutamicum towards higher growth rates on glucose minimal medium

    doi: 10.1038/s41598-017-17014-9

    Figure Lengend Snippet: Adaptive evolution experiment. ( A ) C. glutamicum ATCC 13032 and its prophage-free variant MB001 were adaptively evolved towards increased growth rates on glucose minimal medium (CGXII medium with 2% (w/v) glucose). Overall, six independent cell lines for each strain were evolved by repetitive batch cultivation using 48-well plates incubated at 30 °C (experimental details are given in the material and methods section). ( B ) Time line of the ALE experiment. Highlighted are the samples that were further analyzed by genome sequencing. Samples of the second reproduction of the experiment are marked with an asterisk.

    Article Snippet: In this work, we compared C. glutamicum ATCC 13032 wild type strain and its prophage-free variant MB001 in a long-term evolution experiment on glucose minimal medium.

    Techniques: Variant Assay, Incubation, Sequencing

    Impact of key mutations on the fitness in glucose minimal medium ( A ) Key mutations identified in pyk , fruK and corA (see Table 2 ) were introduced into the parental wild type strain C. glutamicum ATCC 13032 and investigated regarding their impact on growth in CGXII minimal medium containing 2% (w/v) glucose. Each graph is showing the average data of three biological replicates including standard deviation. ( B ). Selected mutations, as found in UBm, were step-wise introduced into the wild type strain and their impact on the growth on glucose containing minimal media was examined for three biological replicates. ( C ) The growth of the wild type, UBm and the single mutant strains (WT_corA_Q307 Δ12 bp) were compared on CGXII minimal medium with 2% (w/v) glucose and different MgSO 4 concentrations (0.05 mM, 1 mM and 10 mM). Pre-cultivations were performed as described in the material and methods part. Each curve represents three biological replicates including standard deviation.

    Journal: Scientific Reports

    Article Title: Adaptive laboratory evolution of Corynebacterium glutamicum towards higher growth rates on glucose minimal medium

    doi: 10.1038/s41598-017-17014-9

    Figure Lengend Snippet: Impact of key mutations on the fitness in glucose minimal medium ( A ) Key mutations identified in pyk , fruK and corA (see Table 2 ) were introduced into the parental wild type strain C. glutamicum ATCC 13032 and investigated regarding their impact on growth in CGXII minimal medium containing 2% (w/v) glucose. Each graph is showing the average data of three biological replicates including standard deviation. ( B ). Selected mutations, as found in UBm, were step-wise introduced into the wild type strain and their impact on the growth on glucose containing minimal media was examined for three biological replicates. ( C ) The growth of the wild type, UBm and the single mutant strains (WT_corA_Q307 Δ12 bp) were compared on CGXII minimal medium with 2% (w/v) glucose and different MgSO 4 concentrations (0.05 mM, 1 mM and 10 mM). Pre-cultivations were performed as described in the material and methods part. Each curve represents three biological replicates including standard deviation.

    Article Snippet: In this work, we compared C. glutamicum ATCC 13032 wild type strain and its prophage-free variant MB001 in a long-term evolution experiment on glucose minimal medium.

    Techniques: Standard Deviation, Mutagenesis

    Competitive growth experiment of C. glutamicum ATCC 13032 and MB001. C. glutamicum ATCC 13032 and MB001 strains were both labelled vice versa with a genomically encoded yellow or far red fluorescent protein (eYFP and E2-crimson) 19 . The resulting strains were compared vice versa in a competitive growth study on CGXII minimal medium containing 2% (w/v) glucose. Cells were serially transferred to fresh medium after 24 h. Production of the fluorescent proteins was induced by adding IPTG in a final concentration of 0.5 mM six hours before samples for flow cytometry analysis were taken. Strains from the co-cultivation experiment were discriminated by flow cytometry determining the fraction of eYFP and E2-Crimson positive cells.

    Journal: Scientific Reports

    Article Title: Adaptive laboratory evolution of Corynebacterium glutamicum towards higher growth rates on glucose minimal medium

    doi: 10.1038/s41598-017-17014-9

    Figure Lengend Snippet: Competitive growth experiment of C. glutamicum ATCC 13032 and MB001. C. glutamicum ATCC 13032 and MB001 strains were both labelled vice versa with a genomically encoded yellow or far red fluorescent protein (eYFP and E2-crimson) 19 . The resulting strains were compared vice versa in a competitive growth study on CGXII minimal medium containing 2% (w/v) glucose. Cells were serially transferred to fresh medium after 24 h. Production of the fluorescent proteins was induced by adding IPTG in a final concentration of 0.5 mM six hours before samples for flow cytometry analysis were taken. Strains from the co-cultivation experiment were discriminated by flow cytometry determining the fraction of eYFP and E2-Crimson positive cells.

    Article Snippet: In this work, we compared C. glutamicum ATCC 13032 wild type strain and its prophage-free variant MB001 in a long-term evolution experiment on glucose minimal medium.

    Techniques: Concentration Assay, Flow Cytometry, Cytometry

    Detachment of THP-1 cells infected with C. ulcerans . Cells infected with non-pathogenic C. glutamicum or pathogenic C. ulcerans at an MOI of 10 and 1 were analyzed microscopically at 2, 8 and 20 h post-infection, untreated cells served as negative

    Journal: Virulence

    Article Title: The killing of macrophages by Corynebacterium ulcerans

    doi: 10.1080/21505594.2015.1125068

    Figure Lengend Snippet: Detachment of THP-1 cells infected with C. ulcerans . Cells infected with non-pathogenic C. glutamicum or pathogenic C. ulcerans at an MOI of 10 and 1 were analyzed microscopically at 2, 8 and 20 h post-infection, untreated cells served as negative

    Article Snippet: In contrast to non-pathogenic C. glutamicum strain ATCC 13032, the C. ulcerans strains were able to multiply within macrophages at least 8 hours post-infection and were able to delay phagolysosome formation, which might be an effective mechanism for immune evasion and spreading of C. ulcerans within the body supporting the establishment and progress of infections.

    Techniques: Infection

    NF-κB activation in THP1-Blue™ NF-κB reporter cells after C. ulcerans infection. THP1-Blue™ NF-κB cells were incubated for 20 h with ( A ) viable and ( B ) UV-killed bacteria of the non-pathogenic C. glutamicum

    Journal: Virulence

    Article Title: The killing of macrophages by Corynebacterium ulcerans

    doi: 10.1080/21505594.2015.1125068

    Figure Lengend Snippet: NF-κB activation in THP1-Blue™ NF-κB reporter cells after C. ulcerans infection. THP1-Blue™ NF-κB cells were incubated for 20 h with ( A ) viable and ( B ) UV-killed bacteria of the non-pathogenic C. glutamicum

    Article Snippet: In contrast to non-pathogenic C. glutamicum strain ATCC 13032, the C. ulcerans strains were able to multiply within macrophages at least 8 hours post-infection and were able to delay phagolysosome formation, which might be an effective mechanism for immune evasion and spreading of C. ulcerans within the body supporting the establishment and progress of infections.

    Techniques: Activation Assay, Infection, Incubation

    7-AAD staining and FACS analysis of THP-1 cells infected with C. ulcerans . THP-1 cells were infected with the non-pathogenic C. glutamicum ATCC 13032 and pathogenic C. ulcerans strains 809 and BR-AD22 at MOI 50, uninfected cells were carried along as

    Journal: Virulence

    Article Title: The killing of macrophages by Corynebacterium ulcerans

    doi: 10.1080/21505594.2015.1125068

    Figure Lengend Snippet: 7-AAD staining and FACS analysis of THP-1 cells infected with C. ulcerans . THP-1 cells were infected with the non-pathogenic C. glutamicum ATCC 13032 and pathogenic C. ulcerans strains 809 and BR-AD22 at MOI 50, uninfected cells were carried along as

    Article Snippet: In contrast to non-pathogenic C. glutamicum strain ATCC 13032, the C. ulcerans strains were able to multiply within macrophages at least 8 hours post-infection and were able to delay phagolysosome formation, which might be an effective mechanism for immune evasion and spreading of C. ulcerans within the body supporting the establishment and progress of infections.

    Techniques: Staining, FACS, Infection

    Fluorescence microscopy of C. ulcerans and THP-1 cells. THP-1 cells were infected with C. glutamicum ATCC 13032 pEPR1p45 gfp, C. ulcerans 809 pEPR1p45 gfp and C. ulcerans BR-AD22 pEPR1p45 gfp at an MOI of 10 for 30 min. Extracellular bacteria were

    Journal: Virulence

    Article Title: The killing of macrophages by Corynebacterium ulcerans

    doi: 10.1080/21505594.2015.1125068

    Figure Lengend Snippet: Fluorescence microscopy of C. ulcerans and THP-1 cells. THP-1 cells were infected with C. glutamicum ATCC 13032 pEPR1p45 gfp, C. ulcerans 809 pEPR1p45 gfp and C. ulcerans BR-AD22 pEPR1p45 gfp at an MOI of 10 for 30 min. Extracellular bacteria were

    Article Snippet: In contrast to non-pathogenic C. glutamicum strain ATCC 13032, the C. ulcerans strains were able to multiply within macrophages at least 8 hours post-infection and were able to delay phagolysosome formation, which might be an effective mechanism for immune evasion and spreading of C. ulcerans within the body supporting the establishment and progress of infections.

    Techniques: Fluorescence, Microscopy, Infection

    Labeling and tracking of acidic organelles in THP-1 cells infected with C. ulcerans . THP-1 cells were incubated with LysoTracker ® Red DND-99 for 120 min before cells were infected with C. glutamicum ATCC 13032 pEPR1p45 gfp, C. ulcerans

    Journal: Virulence

    Article Title: The killing of macrophages by Corynebacterium ulcerans

    doi: 10.1080/21505594.2015.1125068

    Figure Lengend Snippet: Labeling and tracking of acidic organelles in THP-1 cells infected with C. ulcerans . THP-1 cells were incubated with LysoTracker ® Red DND-99 for 120 min before cells were infected with C. glutamicum ATCC 13032 pEPR1p45 gfp, C. ulcerans

    Article Snippet: In contrast to non-pathogenic C. glutamicum strain ATCC 13032, the C. ulcerans strains were able to multiply within macrophages at least 8 hours post-infection and were able to delay phagolysosome formation, which might be an effective mechanism for immune evasion and spreading of C. ulcerans within the body supporting the establishment and progress of infections.

    Techniques: Labeling, Infection, Incubation

    Growth of the gapN -expressing strain. Wild-type C. glutamicum (•), the Δ gapB strain (▪), the Δ gapAB strain (♦), strain GPN (▴), and a suppressor mutant from strain GPN (▵) were cultivated in 5 ml

    Journal: Applied and Environmental Microbiology

    Article Title: Engineering of Corynebacterium glutamicum with an NADPH-Generating Glycolytic Pathway for l-Lysine Production ▿

    doi: 10.1128/AEM.01464-10

    Figure Lengend Snippet: Growth of the gapN -expressing strain. Wild-type C. glutamicum (•), the Δ gapB strain (▪), the Δ gapAB strain (♦), strain GPN (▴), and a suppressor mutant from strain GPN (▵) were cultivated in 5 ml

    Article Snippet: The wild-type C. glutamicum strain ATCC 13032 was used in this study.

    Techniques: Expressing, Mutagenesis

    Growth of the gapN -expressing strain and generation of a spontaneous mutant from the recombinant strain. Wild type C. glutamicum (•), the Δ gapB strain (▪), the Δ gapAB strain (♦), strain GPN (▴), and suppressor

    Journal: Applied and Environmental Microbiology

    Article Title: Engineering of Corynebacterium glutamicum with an NADPH-Generating Glycolytic Pathway for l-Lysine Production ▿

    doi: 10.1128/AEM.01464-10

    Figure Lengend Snippet: Growth of the gapN -expressing strain and generation of a spontaneous mutant from the recombinant strain. Wild type C. glutamicum (•), the Δ gapB strain (▪), the Δ gapAB strain (♦), strain GPN (▴), and suppressor

    Article Snippet: The wild-type C. glutamicum strain ATCC 13032 was used in this study.

    Techniques: Expressing, Mutagenesis, Recombinant

    l- Lysine production and growth levels of C. glutamicum strains on various sugars. Wild type C. glutamicum , the Δ gapB strain, strain GPN, suppressor mutant RE2, and strain RE2A were cultivated in 5 ml of LFG1 medium supplemented with 3%

    Journal: Applied and Environmental Microbiology

    Article Title: Engineering of Corynebacterium glutamicum with an NADPH-Generating Glycolytic Pathway for l-Lysine Production ▿

    doi: 10.1128/AEM.01464-10

    Figure Lengend Snippet: l- Lysine production and growth levels of C. glutamicum strains on various sugars. Wild type C. glutamicum , the Δ gapB strain, strain GPN, suppressor mutant RE2, and strain RE2A were cultivated in 5 ml of LFG1 medium supplemented with 3%

    Article Snippet: The wild-type C. glutamicum strain ATCC 13032 was used in this study.

    Techniques: Mutagenesis

    Comparative transcriptome analysis of l -lactate- and pyruvate-grown C. glutamicum ATCC 13032.

    Journal:

    Article Title: Characterization of a Corynebacterium glutamicum Lactate Utilization Operon Induced during Temperature-Triggered Glutamate Production †

    doi: 10.1128/AEM.71.10.5920-5928.2005

    Figure Lengend Snippet: Comparative transcriptome analysis of l -lactate- and pyruvate-grown C. glutamicum ATCC 13032.

    Article Snippet: To study the function of the putative lactate utilization genes, the genetically tractable wild-type strain C. glutamicum ATCC 13032 was employed ( ).

    Techniques:

    Properties and carbon source-dependent regulation of quinone-dependent l -lactate dehydrogenase from C. glutamicum ATCC 13032.

    Journal:

    Article Title: Characterization of a Corynebacterium glutamicum Lactate Utilization Operon Induced during Temperature-Triggered Glutamate Production †

    doi: 10.1128/AEM.71.10.5920-5928.2005

    Figure Lengend Snippet: Properties and carbon source-dependent regulation of quinone-dependent l -lactate dehydrogenase from C. glutamicum ATCC 13032.

    Article Snippet: To study the function of the putative lactate utilization genes, the genetically tractable wild-type strain C. glutamicum ATCC 13032 was employed ( ).

    Techniques:

    Point mutation and gene insertion mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. a Schematic depicting the procedure for generating point mutations. The 5′ end of LR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the left arm. The point mutation site was designed in the primers FR and RF. CF and CR are primers for PCR validation of editing efficiency. The SEQ primer is used for sequencing. b Schematic depicting the procedure for gene insertion. The 5′ end of the LR contains a 10 bp overhang region of the 5′ end of GFP. The 5′ end of GFPF contains a 10 bp overhang region of the 3′ end of the left arm. The 5′ end of GFPR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the GFP arm. c The point mutation mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. The mutation efficiency was 6/6 and was confirmed by sequencing. d Gene insertion mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. The porB gene was replaced by the gfp gene. The efficiency for gfp insertion was 8/12, confirmed by PCR and sequencing

    Journal: Microbial Cell Factories

    Article Title: Efficient gene editing in Corynebacterium glutamicum using the CRISPR/Cas9 system

    doi: 10.1186/s12934-017-0814-6

    Figure Lengend Snippet: Point mutation and gene insertion mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. a Schematic depicting the procedure for generating point mutations. The 5′ end of LR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the left arm. The point mutation site was designed in the primers FR and RF. CF and CR are primers for PCR validation of editing efficiency. The SEQ primer is used for sequencing. b Schematic depicting the procedure for gene insertion. The 5′ end of the LR contains a 10 bp overhang region of the 5′ end of GFP. The 5′ end of GFPF contains a 10 bp overhang region of the 3′ end of the left arm. The 5′ end of GFPR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the GFP arm. c The point mutation mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. The mutation efficiency was 6/6 and was confirmed by sequencing. d Gene insertion mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. The porB gene was replaced by the gfp gene. The efficiency for gfp insertion was 8/12, confirmed by PCR and sequencing

    Article Snippet: The resulting plasmid, pXMJ19-EGFP, was introduced into the four gene-deleted C. glutamicum ATCC 13032 strains.

    Techniques: Mutagenesis, CRISPR, Polymerase Chain Reaction, Sequencing

    CRISPR/Cas9-mediated genome editing in C. glutamicum ATCC 13032 and C. glutamicum CGMCC1.15647. a Schematic depicting editing procedures. The left and right arms are regions from the targeted gene and are amplified by PCR from C. glutamicum genomic DNA. LF and LR primers are used to amplify the left arm, and RF and RR primers are used to amplify the right arm. For Gibson assembly, the 5′ end of LF contains a 20 bp overhang region of the 5′ end of the Bgl II site from the pFST plasmid. The 5′ end of LR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the left arm. The 5′ end of RR contains a 20 bp overhang region of the 3′ end of the Bgl II site from the pFST plasmid. CF and CR are primers for PCR validation of editing efficiency. The SEQ primer is used for sequencing. b The CRISPR/Cas9 system mediated disruption of the porB gene in C. glutamicum ATCC 13032. The editing efficiency was 18/18. The lane ‘ck’ is the PCR product from the wild-type strain. These results were confirmed by sequencing. c The CRISPR/Cas9 system mediated disruption of the proB gene in C. glutamicum CGMCC1.15647. The editing efficiency was 16/16

    Journal: Microbial Cell Factories

    Article Title: Efficient gene editing in Corynebacterium glutamicum using the CRISPR/Cas9 system

    doi: 10.1186/s12934-017-0814-6

    Figure Lengend Snippet: CRISPR/Cas9-mediated genome editing in C. glutamicum ATCC 13032 and C. glutamicum CGMCC1.15647. a Schematic depicting editing procedures. The left and right arms are regions from the targeted gene and are amplified by PCR from C. glutamicum genomic DNA. LF and LR primers are used to amplify the left arm, and RF and RR primers are used to amplify the right arm. For Gibson assembly, the 5′ end of LF contains a 20 bp overhang region of the 5′ end of the Bgl II site from the pFST plasmid. The 5′ end of LR contains a 10 bp overhang region of the 5′ end of the right arm. The 5′ end of RF contains a 10 bp overhang region of the 3′ end of the left arm. The 5′ end of RR contains a 20 bp overhang region of the 3′ end of the Bgl II site from the pFST plasmid. CF and CR are primers for PCR validation of editing efficiency. The SEQ primer is used for sequencing. b The CRISPR/Cas9 system mediated disruption of the porB gene in C. glutamicum ATCC 13032. The editing efficiency was 18/18. The lane ‘ck’ is the PCR product from the wild-type strain. These results were confirmed by sequencing. c The CRISPR/Cas9 system mediated disruption of the proB gene in C. glutamicum CGMCC1.15647. The editing efficiency was 16/16

    Article Snippet: The resulting plasmid, pXMJ19-EGFP, was introduced into the four gene-deleted C. glutamicum ATCC 13032 strains.

    Techniques: CRISPR, Amplification, Polymerase Chain Reaction, Plasmid Preparation, Sequencing

    Genome editing mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. a Schematic depicting editing procedures, the PAM site and gene length. b Disruption of the mepA gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 2/15. The lane ‘ck’ is the PCR product from the wild-type strain. c Disruption of the clpX gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 5/16. d Disruption of the Ncgl0911 gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 4/15

    Journal: Microbial Cell Factories

    Article Title: Efficient gene editing in Corynebacterium glutamicum using the CRISPR/Cas9 system

    doi: 10.1186/s12934-017-0814-6

    Figure Lengend Snippet: Genome editing mediated by CRISPR/Cas9 in C. glutamicum ATCC 13032. a Schematic depicting editing procedures, the PAM site and gene length. b Disruption of the mepA gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 2/15. The lane ‘ck’ is the PCR product from the wild-type strain. c Disruption of the clpX gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 5/16. d Disruption of the Ncgl0911 gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032. The editing efficiency was 4/15

    Article Snippet: The resulting plasmid, pXMJ19-EGFP, was introduced into the four gene-deleted C. glutamicum ATCC 13032 strains.

    Techniques: CRISPR, Polymerase Chain Reaction

    Evaluation of editing efficiency with different arm sizes. a Design of HDarms of various sizes (600, 300, 100 bp). Both sides of the HDarm contain a 20 bp overhang region of the Bgl II site from the pFST plasmid. b Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 600 bp HDarm. The editing efficiency was 10/12, the lane ‘ck’ is the PCR product from the wild-type strain. c Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 300 bp HDarm. The editing efficiency was 10/12. d Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 100 bp HDarm. The editing efficiency was 2/12

    Journal: Microbial Cell Factories

    Article Title: Efficient gene editing in Corynebacterium glutamicum using the CRISPR/Cas9 system

    doi: 10.1186/s12934-017-0814-6

    Figure Lengend Snippet: Evaluation of editing efficiency with different arm sizes. a Design of HDarms of various sizes (600, 300, 100 bp). Both sides of the HDarm contain a 20 bp overhang region of the Bgl II site from the pFST plasmid. b Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 600 bp HDarm. The editing efficiency was 10/12, the lane ‘ck’ is the PCR product from the wild-type strain. c Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 300 bp HDarm. The editing efficiency was 10/12. d Disruption of the porB gene mediated by the CRISPR/Cas9 system in C. glutamicum ATCC 13032 with a 100 bp HDarm. The editing efficiency was 2/12

    Article Snippet: The resulting plasmid, pXMJ19-EGFP, was introduced into the four gene-deleted C. glutamicum ATCC 13032 strains.

    Techniques: Plasmid Preparation, CRISPR, Polymerase Chain Reaction

    Live cell imaging of L-valine production strains using microfluidic monolayer cultivation chambers . ( A ) Illustration of the microfluidic cultivation chambers. The system consists of several arrays of picoliter sized monolayer cultivation chambers. ( B ) Fluorescence emission of three entire microcolonies (average eYFP signal per colony area) of the ΔaceE sensor strain (triangles) and the ΔaceE Δpqo Δpgi sensor strain (diamonds) over time. Fluorescence was measured every 2.5 h. ( C ) Growth (t 1 –t 2 ) and production phase (t 3 –t 5 ) of isogenic microcolonies of the ΔaceE sensor strain (upper row) and the ΔaceE Δpqo Δpgi sensor strain (lower row). ( D ) Histograms illustrating fluorescence distribution within a representative microcolony of the ΔaceE sensor strain (left) and the ΔaceE Δpqo Δpgi sensor strain (right). The eYFP signal of single cells was measured at t = 19 h (red), t = 26 h (green), t = 34 h (purple), and t = 46 h (blue). Average fluorescence values are indicated above the respective peaks. All cultivations were performed in microfluidic chambers shown in (A) in CGXII minimal medium containing 154 mM acetate and 222 mM glucose during growth phase or CGXII with 222 mM glucose during the production phase, respectively.

    Journal: PLoS ONE

    Article Title: Application of a Genetically Encoded Biosensor for Live Cell Imaging of L-Valine Production in Pyruvate Dehydrogenase Complex-Deficient Corynebacterium glutamicum Strains

    doi: 10.1371/journal.pone.0085731

    Figure Lengend Snippet: Live cell imaging of L-valine production strains using microfluidic monolayer cultivation chambers . ( A ) Illustration of the microfluidic cultivation chambers. The system consists of several arrays of picoliter sized monolayer cultivation chambers. ( B ) Fluorescence emission of three entire microcolonies (average eYFP signal per colony area) of the ΔaceE sensor strain (triangles) and the ΔaceE Δpqo Δpgi sensor strain (diamonds) over time. Fluorescence was measured every 2.5 h. ( C ) Growth (t 1 –t 2 ) and production phase (t 3 –t 5 ) of isogenic microcolonies of the ΔaceE sensor strain (upper row) and the ΔaceE Δpqo Δpgi sensor strain (lower row). ( D ) Histograms illustrating fluorescence distribution within a representative microcolony of the ΔaceE sensor strain (left) and the ΔaceE Δpqo Δpgi sensor strain (right). The eYFP signal of single cells was measured at t = 19 h (red), t = 26 h (green), t = 34 h (purple), and t = 46 h (blue). Average fluorescence values are indicated above the respective peaks. All cultivations were performed in microfluidic chambers shown in (A) in CGXII minimal medium containing 154 mM acetate and 222 mM glucose during growth phase or CGXII with 222 mM glucose during the production phase, respectively.

    Article Snippet: Occurrence of non-fluorescent cells during the production phase of the C. glutamicum ATCC 13032 ΔaceE Δpqo Δpgi sensor strain. (WMV) Click here for additional data file.

    Techniques: Live Cell Imaging, Fluorescence

    Occurrence of non-fluorescent cells during the production phase. ( A ) Microcolony and lineage tree of the ΔaceE Δpqo Δpgi sensor strain. Different types of non-fluorescent cells are illustrated in B. ( B ) (I+II) Lysing cells and (III) dormant/or dead cell, which do not switch from growth to production. (IV) Leaky cell that shows decreasing fluorescence signal over time, potentially caused by a permeabilized cell membrane. (V * ) Cells showing slow growth, but no production. Images marked with an asterisk show cells of another microcolony of the ΔaceE Δpqo Δpgi sensor strain, not shown in this figure.

    Journal: PLoS ONE

    Article Title: Application of a Genetically Encoded Biosensor for Live Cell Imaging of L-Valine Production in Pyruvate Dehydrogenase Complex-Deficient Corynebacterium glutamicum Strains

    doi: 10.1371/journal.pone.0085731

    Figure Lengend Snippet: Occurrence of non-fluorescent cells during the production phase. ( A ) Microcolony and lineage tree of the ΔaceE Δpqo Δpgi sensor strain. Different types of non-fluorescent cells are illustrated in B. ( B ) (I+II) Lysing cells and (III) dormant/or dead cell, which do not switch from growth to production. (IV) Leaky cell that shows decreasing fluorescence signal over time, potentially caused by a permeabilized cell membrane. (V * ) Cells showing slow growth, but no production. Images marked with an asterisk show cells of another microcolony of the ΔaceE Δpqo Δpgi sensor strain, not shown in this figure.

    Article Snippet: Occurrence of non-fluorescent cells during the production phase of the C. glutamicum ATCC 13032 ΔaceE Δpqo Δpgi sensor strain. (WMV) Click here for additional data file.

    Techniques: Fluorescence

    Levels of select transcripts in the evolved strains C . glutamicum ΔAPE6937 (A) and C . glutamicum ΔAPE6937R42 (B) compared with those of the parent strain C . glutamicum ΔAPE at 54 h in shake flasks. Abundances of each transcript in ΔAPE6937 or ΔAPE6937R42, determined using qRT-PCR, were normalised relative to levels of the same transcript in the parental strain.

    Journal: BMC Biotechnology

    Article Title: Metabolic evolution of Corynebacterium glutamicum for increased production of L-ornithine

    doi: 10.1186/1472-6750-13-47

    Figure Lengend Snippet: Levels of select transcripts in the evolved strains C . glutamicum ΔAPE6937 (A) and C . glutamicum ΔAPE6937R42 (B) compared with those of the parent strain C . glutamicum ΔAPE at 54 h in shake flasks. Abundances of each transcript in ΔAPE6937 or ΔAPE6937R42, determined using qRT-PCR, were normalised relative to levels of the same transcript in the parental strain.

    Article Snippet: Hwang et al. reported that co-overexpression of argCJBD in a triple-gene knockout strain C. glutamicum ATCC 13032 (ΔargFΔargRΔproB ) resulted in a cellular L-ornithine content of 16.49 mg/g DCW and a concentration of L-ornithine in the culture medium of 179.14 mg/L [ ].

    Techniques: Quantitative RT-PCR

    Scheme of reconstruction of cellobiose-utilizing and xylose-utilizing pathway in C. glutamicum ATCC 13032. C. glutamicum wild-type is not able to utilize cellobiose and xylose as sole carbon source ( left ). No genes for xylose isomerase, cellobiose transporters, and β-glucosidase are annotated (shown as no arrow ). Through metabolic engineering and adaptive evolution of C. glutamicum strains ( right ), the cells were able to utilize cellobiose and xylose. Extracellular cellobiose was transported and intracellular β-glucosidase encoded by the gh1 - 1 gene hydrolyzed intracellular cellobiose to glucose, which further was metabolized by glucokinase (Glk) into glycolysis. Xylose metabolic pathway consists of heterologous xylose isomerase ( xylA from E. coli ) and additional xylulose kinase ( xylB from E. coli ). Transporter of Gxf1 ( Candida intermedia ) and Sut1 ( Pichia stipitis ), respectively was introduced as a xylose transporter

    Journal: Microbial Cell Factories

    Article Title: Adaptive evolution and metabolic engineering of a cellobiose- and xylose- negative Corynebacterium glutamicum that co-utilizes cellobiose and xylose

    doi: 10.1186/s12934-016-0420-z

    Figure Lengend Snippet: Scheme of reconstruction of cellobiose-utilizing and xylose-utilizing pathway in C. glutamicum ATCC 13032. C. glutamicum wild-type is not able to utilize cellobiose and xylose as sole carbon source ( left ). No genes for xylose isomerase, cellobiose transporters, and β-glucosidase are annotated (shown as no arrow ). Through metabolic engineering and adaptive evolution of C. glutamicum strains ( right ), the cells were able to utilize cellobiose and xylose. Extracellular cellobiose was transported and intracellular β-glucosidase encoded by the gh1 - 1 gene hydrolyzed intracellular cellobiose to glucose, which further was metabolized by glucokinase (Glk) into glycolysis. Xylose metabolic pathway consists of heterologous xylose isomerase ( xylA from E. coli ) and additional xylulose kinase ( xylB from E. coli ). Transporter of Gxf1 ( Candida intermedia ) and Sut1 ( Pichia stipitis ), respectively was introduced as a xylose transporter

    Article Snippet: Finally, we obtained Cg -Cello01(evo) and Cg -Cello02(evo) strains as cellobiose-utilizing C. glutamicum ATCC 13032 derivative strains.

    Techniques:

    Schematic representation of genes involved in arsenic resistance in C. glutamicum ATCC 13032 (A) and in other bacteria (B). (A) Operons ars1 and ars2 are indicated. Arrows represent open reading frames. Dashed boxes indicate the probes used for the cloning

    Journal:

    Article Title: Analysis of Genes Involved in Arsenic Resistance in Corynebacterium glutamicum ATCC 13032 †

    doi: 10.1128/AEM.71.10.6206-6215.2005

    Figure Lengend Snippet: Schematic representation of genes involved in arsenic resistance in C. glutamicum ATCC 13032 (A) and in other bacteria (B). (A) Operons ars1 and ars2 are indicated. Arrows represent open reading frames. Dashed boxes indicate the probes used for the cloning

    Article Snippet: Primers for PCR were generated by using the C. glutamicum strain ATCC 13032 genome sequence and are listed in Table S1 in the supplemental material.

    Techniques: Clone Assay

    Ratio/intensity ( m / a ) plot deduced from DNA microarray hybridizations comparing the transcriptome of the dtxR mutant C. glutamicum IB2103 with that of the wild-type strain C. glutamicum ATCC 13032. The analyzed strains were cultivated in low-iron CGXII medium to mid-exponential growth phase. Then, 10 mg/l FeSO 4 was added to the medium, and the cultures were incubated for further 15 min. Subsequently, total RNA was isolated from two biological replicates and used for DNA microarray hybridizations including label swapping. Genes showing enhanced expression in the dtxR mutant C. glutamicum IB2103 are marked by black diamonds; genes with decreased expression are indicated by black triangles; genes without differential expression pattern are shown by gray spots. Differentially expressed genes later on assigned to the DtxR regulon of C. glutamicum are named by their identifiers. Genes were regarded as being differentially expressed with a -values equal or greater than 7.0 and m -values equal or greater than 1 (up-regulation) or equal or smaller than -1 (down-regulation).

    Journal: BMC Genomics

    Article Title: The DtxR protein acting as dual transcriptional regulator directs a global regulatory network involved in iron metabolism of Corynebacterium glutamicum

    doi: 10.1186/1471-2164-7-21

    Figure Lengend Snippet: Ratio/intensity ( m / a ) plot deduced from DNA microarray hybridizations comparing the transcriptome of the dtxR mutant C. glutamicum IB2103 with that of the wild-type strain C. glutamicum ATCC 13032. The analyzed strains were cultivated in low-iron CGXII medium to mid-exponential growth phase. Then, 10 mg/l FeSO 4 was added to the medium, and the cultures were incubated for further 15 min. Subsequently, total RNA was isolated from two biological replicates and used for DNA microarray hybridizations including label swapping. Genes showing enhanced expression in the dtxR mutant C. glutamicum IB2103 are marked by black diamonds; genes with decreased expression are indicated by black triangles; genes without differential expression pattern are shown by gray spots. Differentially expressed genes later on assigned to the DtxR regulon of C. glutamicum are named by their identifiers. Genes were regarded as being differentially expressed with a -values equal or greater than 7.0 and m -values equal or greater than 1 (up-regulation) or equal or smaller than -1 (down-regulation).

    Article Snippet: Bacterial strains and growth conditions Wild-type strain C. glutamicum ATCC 13032 (American Type Culture Collection, Manassas, VA) was used to characterize the DtxR regulon.

    Techniques: Microarray, Mutagenesis, Incubation, Isolation, Expressing

    Growth curves of the wild-type strain C. glutamicum ATCC 13032 and the dtxR mutant C. glutamicum IB2103. The strains and plasmid-carrying derivatives were cultivated in CGXII medium and in low-iron CGXII medium, respectively. Growth was monitored with a nephelometer and is shown as relative turbidity. Values are means of measurements of six biological replicates. Abbreviations: A, C. glutamicum ATCC 13032 in CGXII medium; B, C. glutamicum IB2103 in CGXII medium; C, C. glutamicum ATCC 13032 in low-iron CGXII medium; D, C. glutamicum IB2103 in low-iron CGXII medium; E, C. glutamicum ATCC 13032 (pIB4000) in CGXII medium; F, C. glutamicum IB2103 (pIB4000) in CGXII medium; G, C. glutamicum ATCC 13032 (pEC-XK99E) in CGXII medium.

    Journal: BMC Genomics

    Article Title: The DtxR protein acting as dual transcriptional regulator directs a global regulatory network involved in iron metabolism of Corynebacterium glutamicum

    doi: 10.1186/1471-2164-7-21

    Figure Lengend Snippet: Growth curves of the wild-type strain C. glutamicum ATCC 13032 and the dtxR mutant C. glutamicum IB2103. The strains and plasmid-carrying derivatives were cultivated in CGXII medium and in low-iron CGXII medium, respectively. Growth was monitored with a nephelometer and is shown as relative turbidity. Values are means of measurements of six biological replicates. Abbreviations: A, C. glutamicum ATCC 13032 in CGXII medium; B, C. glutamicum IB2103 in CGXII medium; C, C. glutamicum ATCC 13032 in low-iron CGXII medium; D, C. glutamicum IB2103 in low-iron CGXII medium; E, C. glutamicum ATCC 13032 (pIB4000) in CGXII medium; F, C. glutamicum IB2103 (pIB4000) in CGXII medium; G, C. glutamicum ATCC 13032 (pEC-XK99E) in CGXII medium.

    Article Snippet: Bacterial strains and growth conditions Wild-type strain C. glutamicum ATCC 13032 (American Type Culture Collection, Manassas, VA) was used to characterize the DtxR regulon.

    Techniques: Mutagenesis, Plasmid Preparation

    Analysis of the in vivo OdhI phosphorylation status in cells grown for 24 h in BHI medium with 4% (w/v) glucose. A. Western blot analysis of cell-free protein extracts (20 μg each) of the indicated C. glutamicum strains with OdhI antibodies. The experiment was performed in triplicate (a, b, c). The upper band in the Western blot represents singly or doubly phosphorylated OdhI, the lower band unphosphorylated OdhI ( Schultz et al. , 2007 ). The percentages (mean values with standard deviation) of unphosphorylated OdhI (black bars) to phosphorylated OdhI (grey bars) were calculated by densitometry. B. Two-dimensional polyacrylamide gel electrophoresis of cell-free protein extracts (300 μg each) of the indicated C. glutamicum strains. In the left part of the figure, sections of three 2D gels prepared with extracts of three independent cultures of the respective strain are shown. The three spots representing unphosphorylated, monophosphorylated and presumably diphosphorylated OdhI are circled and labelled with 0, 1 and 2 respectively. The relative amounts of the three OdhI forms were calculated by densitometric analysis of the OdhI spots. Normalization was performed using the spot labelled X that represents adenylate kinase (cg0648), a protein whose intracellular level is apparently not influenced by the deletion of STPKs or Ppp. The percentage of unphosphorylated OdhI is shown with black bars, that of monophosphorylated OdhI with grey bars, and that of presumably diphosphorylated OdhI by white bars. The spot labelled Y represents MenG ( S -adenosylmethionine:2-demethylmenaquinone methyltransferase, cg1055) and is shown because of the small distance to the OdhI spot labelled 2.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Analysis of the in vivo OdhI phosphorylation status in cells grown for 24 h in BHI medium with 4% (w/v) glucose. A. Western blot analysis of cell-free protein extracts (20 μg each) of the indicated C. glutamicum strains with OdhI antibodies. The experiment was performed in triplicate (a, b, c). The upper band in the Western blot represents singly or doubly phosphorylated OdhI, the lower band unphosphorylated OdhI ( Schultz et al. , 2007 ). The percentages (mean values with standard deviation) of unphosphorylated OdhI (black bars) to phosphorylated OdhI (grey bars) were calculated by densitometry. B. Two-dimensional polyacrylamide gel electrophoresis of cell-free protein extracts (300 μg each) of the indicated C. glutamicum strains. In the left part of the figure, sections of three 2D gels prepared with extracts of three independent cultures of the respective strain are shown. The three spots representing unphosphorylated, monophosphorylated and presumably diphosphorylated OdhI are circled and labelled with 0, 1 and 2 respectively. The relative amounts of the three OdhI forms were calculated by densitometric analysis of the OdhI spots. Normalization was performed using the spot labelled X that represents adenylate kinase (cg0648), a protein whose intracellular level is apparently not influenced by the deletion of STPKs or Ppp. The percentage of unphosphorylated OdhI is shown with black bars, that of monophosphorylated OdhI with grey bars, and that of presumably diphosphorylated OdhI by white bars. The spot labelled Y represents MenG ( S -adenosylmethionine:2-demethylmenaquinone methyltransferase, cg1055) and is shown because of the small distance to the OdhI spot labelled 2.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: In Vivo, Western Blot, Standard Deviation, Polyacrylamide Gel Electrophoresis

    Model of STPK-dependent phosphorylation and Ppp-dependent dephosphorylation of OdhI and FtsZ in C. glutamicum .

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Model of STPK-dependent phosphorylation and Ppp-dependent dephosphorylation of OdhI and FtsZ in C. glutamicum .

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: De-Phosphorylation Assay

    Identification of FtsZ as an in vivo substrate of Ppp and as an in vitro substrate for PknA 1−287 , PknB 1−287 and PknL 1−287 . A. Analysis of the FtsZ phosphorylation status in vivo by 2D gel electrophoresis of cell-free extracts of C. glutamicum wild type and C. glutamicum Δ ppp . Both strains were cultivated for 24 h in BHI medium with 4% (w/v) glucose and 300 μg of protein of cell-free extracts was used for separation by 2D-PAGE. For isoelectric focusing, IPG strips (GE Healthcare) with a pH gradient from 4.0 to 5.0 were used, for subsequent SDS-PAGE Excel SDS gradient gels 12–14% (GE Healthcare). In the left part of the figure, sections of three 2D gels prepared from cell extracts of three distinct cultures of the respective strain are shown. The four spots representing presumably unphosphorylated, monophosphorylated, diphosphorylated and triphosphorylated FtsZ are circled and labelled with 0, 1, 2 and 3 respectively. The relative amounts of the four detectable FtsZ isoforms were calculated by densitometric analysis of the FtsZ spots (spot 0, black bars; spot 1, dark grey bars; spot 2, light grey bars; spot 3, white bars). B. Full-length FtsZ (amino acid residues 1–442; 49.7 kDa) containing an N-terminal His tag was overproduced in E. coli and purified by Ni 2+ -chelate affinity chromatography. A sample was subjected to SDS-PAGE and stained with Coomassie brilliant blue. Purified FtsZ was incubated with PknA 1−287 , PknB 1−287 , PknG 1−342 or PknL 1−287 and [γ- 33 P]-ATP and analysed by autoradiography. C. FtsZ phosphopeptides and phosphorylation sites identified by MALDI-TOF-MS and ESI-TOF-MS of tryptic digests of spot 3 in the 2D gel of the Δ ppp mutant and of tryptic digests after in vitro phosphorylation of FtsZ with PknA 1−287 and PknL 1−287 . The FtsZ sequence coverage was between 35% and 45% and did not include all serine and threonine residues present in FtsZ. Unambiguously identified phosphorylation sites are shown in bold and underlined.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Identification of FtsZ as an in vivo substrate of Ppp and as an in vitro substrate for PknA 1−287 , PknB 1−287 and PknL 1−287 . A. Analysis of the FtsZ phosphorylation status in vivo by 2D gel electrophoresis of cell-free extracts of C. glutamicum wild type and C. glutamicum Δ ppp . Both strains were cultivated for 24 h in BHI medium with 4% (w/v) glucose and 300 μg of protein of cell-free extracts was used for separation by 2D-PAGE. For isoelectric focusing, IPG strips (GE Healthcare) with a pH gradient from 4.0 to 5.0 were used, for subsequent SDS-PAGE Excel SDS gradient gels 12–14% (GE Healthcare). In the left part of the figure, sections of three 2D gels prepared from cell extracts of three distinct cultures of the respective strain are shown. The four spots representing presumably unphosphorylated, monophosphorylated, diphosphorylated and triphosphorylated FtsZ are circled and labelled with 0, 1, 2 and 3 respectively. The relative amounts of the four detectable FtsZ isoforms were calculated by densitometric analysis of the FtsZ spots (spot 0, black bars; spot 1, dark grey bars; spot 2, light grey bars; spot 3, white bars). B. Full-length FtsZ (amino acid residues 1–442; 49.7 kDa) containing an N-terminal His tag was overproduced in E. coli and purified by Ni 2+ -chelate affinity chromatography. A sample was subjected to SDS-PAGE and stained with Coomassie brilliant blue. Purified FtsZ was incubated with PknA 1−287 , PknB 1−287 , PknG 1−342 or PknL 1−287 and [γ- 33 P]-ATP and analysed by autoradiography. C. FtsZ phosphopeptides and phosphorylation sites identified by MALDI-TOF-MS and ESI-TOF-MS of tryptic digests of spot 3 in the 2D gel of the Δ ppp mutant and of tryptic digests after in vitro phosphorylation of FtsZ with PknA 1−287 and PknL 1−287 . The FtsZ sequence coverage was between 35% and 45% and did not include all serine and threonine residues present in FtsZ. Unambiguously identified phosphorylation sites are shown in bold and underlined.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: In Vivo, In Vitro, Two-Dimensional Gel Electrophoresis, Electrophoresis, Polyacrylamide Gel Electrophoresis, SDS Page, Purification, Affinity Chromatography, Staining, Incubation, Autoradiography, Mass Spectrometry, Mutagenesis, Sequencing

    Identification of in vivo phosphorylation sites of OdhI. A. Analysis of the in vivo OdhI phosphorylation status by 2D gel electrophoresis of cell-free protein extract of C. glutamicum Δ ppp /pJC1- odhI . The strain was cultivated for 24 h in BHI medium with 4% (w/v) glucose and 300 μg of protein was used for separation by 2D-PAGE. The two spots representing native monophosphorylated and diphosphorylated OdhI are labelled as 1 and 2, respectively, and Strep -tagged monophosphorylated, diphosphorylated and presumably triphosphorylated OdhI are labelled with 1′, 2′ and 3′ respectively. B. MALDI-TOF-MS and MS/MS analysis of tryptically digested OdhI spots 1′ (panel a), 2′ (panel b) and 3′ (panel c) of strain C. glutamicum Δ ppp /pJC1- odhI . Peaks are labelled with their monoisotopic masses. The only phosphopeptide detected was the N-terminal one composed of amino acids 2–19. In the unphoshorylated state, the predicted mass (in the H + form) is 1977.9 Da, in the monophosphorylated state 2057.9 Da, in the diphosphorylated state 2137.9 Da. In panels d and e, MALDI-TOF tandem MS of the 2057.9 Da peptide derived from spot 2′ and of the 2137.8 Da peptide derived from spot 3′ are shown respectively. β-Elimination of phosphoric acid (mass shifts of −18 Da or −36 Da compared to the unphosphorylated fragment after a single or a double β-elimination respectively) is indicated.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Identification of in vivo phosphorylation sites of OdhI. A. Analysis of the in vivo OdhI phosphorylation status by 2D gel electrophoresis of cell-free protein extract of C. glutamicum Δ ppp /pJC1- odhI . The strain was cultivated for 24 h in BHI medium with 4% (w/v) glucose and 300 μg of protein was used for separation by 2D-PAGE. The two spots representing native monophosphorylated and diphosphorylated OdhI are labelled as 1 and 2, respectively, and Strep -tagged monophosphorylated, diphosphorylated and presumably triphosphorylated OdhI are labelled with 1′, 2′ and 3′ respectively. B. MALDI-TOF-MS and MS/MS analysis of tryptically digested OdhI spots 1′ (panel a), 2′ (panel b) and 3′ (panel c) of strain C. glutamicum Δ ppp /pJC1- odhI . Peaks are labelled with their monoisotopic masses. The only phosphopeptide detected was the N-terminal one composed of amino acids 2–19. In the unphoshorylated state, the predicted mass (in the H + form) is 1977.9 Da, in the monophosphorylated state 2057.9 Da, in the diphosphorylated state 2137.9 Da. In panels d and e, MALDI-TOF tandem MS of the 2057.9 Da peptide derived from spot 2′ and of the 2137.8 Da peptide derived from spot 3′ are shown respectively. β-Elimination of phosphoric acid (mass shifts of −18 Da or −36 Da compared to the unphosphorylated fragment after a single or a double β-elimination respectively) is indicated.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: In Vivo, Two-Dimensional Gel Electrophoresis, Electrophoresis, Polyacrylamide Gel Electrophoresis, Mass Spectrometry, Derivative Assay

    In vitro dephosphorylation of phosphorylated OdhI by Ppp. Ppp 1−309 (34.7 kDa) containing an N-terminal decahistidine tag was overproduced in E. coli and purified by Ni 2+ -chelate affinity chromatography. The in vitro dephosphorylation assay was performed as described in Experimental procedures . Lane 1, unphosphorylated OdhI Strep (purified from E. coli BB1553/pAN3K-odhI); lane 2, phosphorylated OdhI Strep (purified from C. glutamicum Δ ppp /pJC1- odhI ); lane 3–8, phosphorylated OdhI Strep 1, 5, 10, 15, 30 and 120 min after addition of the phosphatase Ppp 1−309 . The samples were subjected to SDS-PAGE and stained with Coomassie brilliant blue. The upper band represents singly or doubly phosphorylated OdhI, the lower band unphosphorylated OdhI ( Schultz et al. , 2007 ). The percentage of unphosphorylated (black bars) and phosphorylated OdhI (grey bars) was calculated by densitometric analysis.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: In vitro dephosphorylation of phosphorylated OdhI by Ppp. Ppp 1−309 (34.7 kDa) containing an N-terminal decahistidine tag was overproduced in E. coli and purified by Ni 2+ -chelate affinity chromatography. The in vitro dephosphorylation assay was performed as described in Experimental procedures . Lane 1, unphosphorylated OdhI Strep (purified from E. coli BB1553/pAN3K-odhI); lane 2, phosphorylated OdhI Strep (purified from C. glutamicum Δ ppp /pJC1- odhI ); lane 3–8, phosphorylated OdhI Strep 1, 5, 10, 15, 30 and 120 min after addition of the phosphatase Ppp 1−309 . The samples were subjected to SDS-PAGE and stained with Coomassie brilliant blue. The upper band represents singly or doubly phosphorylated OdhI, the lower band unphosphorylated OdhI ( Schultz et al. , 2007 ). The percentage of unphosphorylated (black bars) and phosphorylated OdhI (grey bars) was calculated by densitometric analysis.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: In Vitro, De-Phosphorylation Assay, Purification, Affinity Chromatography, SDS Page, Staining

    Cell morphology of STPK and Ppp mutants of C. glutamicum growing in BHI medium with 4% (w/v) glucose at 30°C (early exponential phase) was observed. Phase-contrast microscopy pictures are shown and 500 cells of each strain were counted to determine the frequency of cell length. The dotted lines represent the length range from 1.5 to 3 μm which covers 75% of the wild-type cells.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Cell morphology of STPK and Ppp mutants of C. glutamicum growing in BHI medium with 4% (w/v) glucose at 30°C (early exponential phase) was observed. Phase-contrast microscopy pictures are shown and 500 cells of each strain were counted to determine the frequency of cell length. The dotted lines represent the length range from 1.5 to 3 μm which covers 75% of the wild-type cells.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: Microscopy

    Genomic organization of pknA , pknB , pknG , pknL and ppp in C. glutamicum (A) and domain architecture of the corresponding proteins as predicted by PFAM, TMHMM or TMPred (B). KD, kinase domain; TM, transmembrane helix; PP2C, phosphatase domain; PASTA, penicillin-binding protein and serine/threonine kinase-associated domain.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Genomic organization of pknA , pknB , pknG , pknL and ppp in C. glutamicum (A) and domain architecture of the corresponding proteins as predicted by PFAM, TMHMM or TMPred (B). KD, kinase domain; TM, transmembrane helix; PP2C, phosphatase domain; PASTA, penicillin-binding protein and serine/threonine kinase-associated domain.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques: Binding Assay

    Growth of STPK and Ppp mutants of C. glutamicum . A. Growth of different C. glutamicum mutants (black symbols) in comparison with the wild type (white symbols) in BHI medium containing 4% (w/v) glucose. Mean values of triplicate experiments are shown. The final optical densities at 600 nm (OD 600 ) with standard deviations and the maximal growth rates are listed below as well as the P -values from a t -test for pairwise comparisons of the mutants with the wild type. B. Growth on CGXII agar plates containing 100 mM l -glutamine as sole carbon and nitrogen source.

    Journal: Molecular Microbiology

    Article Title: Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases

    doi: 10.1111/j.1365-2958.2009.06897.x

    Figure Lengend Snippet: Growth of STPK and Ppp mutants of C. glutamicum . A. Growth of different C. glutamicum mutants (black symbols) in comparison with the wild type (white symbols) in BHI medium containing 4% (w/v) glucose. Mean values of triplicate experiments are shown. The final optical densities at 600 nm (OD 600 ) with standard deviations and the maximal growth rates are listed below as well as the P -values from a t -test for pairwise comparisons of the mutants with the wild type. B. Growth on CGXII agar plates containing 100 mM l -glutamine as sole carbon and nitrogen source.

    Article Snippet: Reasons for the differing results might be the use of different strains of C. glutamicum (ATCC 13032 and ATCC 13869) or the use of different methods for gene inactivation.

    Techniques:

    Phenotypes of C. glutamicum IMpgm during cultivation in minimal medium with glucose (A, C) or maltose (B, D) Shown are overlays of phase-contrast images with fluorescence images stained with viability stains ( A , B , pictures taken after 7 h of cultivation) or with membrane stain Nile red and DNA stain Hoechst ( C , D , pictures taken after 24 h of cultivation).

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Phenotypes of C. glutamicum IMpgm during cultivation in minimal medium with glucose (A, C) or maltose (B, D) Shown are overlays of phase-contrast images with fluorescence images stained with viability stains ( A , B , pictures taken after 7 h of cultivation) or with membrane stain Nile red and DNA stain Hoechst ( C , D , pictures taken after 24 h of cultivation).

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques: Fluorescence, Staining

    Pgm isoenzymes of C. glutamicum Cell extracts of C. glutamicum WT and C. glutamicum IMpgm were analysed by native PAGE followed by enzyme activity staining ( A ). Total Pgm activity in cell extracts of C. glutamicum WT (filled circles) and C. glutamicum IMpgm (open diamonds) at different concentrations (0.05–20 mM) of glc-1-P ( B ). Data represent mean values of three independent determinations from two independent cultivations and were fitted according to the Michaelis–Menten equation.

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Pgm isoenzymes of C. glutamicum Cell extracts of C. glutamicum WT and C. glutamicum IMpgm were analysed by native PAGE followed by enzyme activity staining ( A ). Total Pgm activity in cell extracts of C. glutamicum WT (filled circles) and C. glutamicum IMpgm (open diamonds) at different concentrations (0.05–20 mM) of glc-1-P ( B ). Data represent mean values of three independent determinations from two independent cultivations and were fitted according to the Michaelis–Menten equation.

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques: Clear Native PAGE, Activity Assay, Staining, Gas Chromatography

    Purification of the main Pgm isoenzyme of C. glutamicum , SDS/PAGE analysis of each step of the purification procedure: ce (crude extract), aec (anion exchange chromatography step wise elution), hic (hydrophobic interaction chromatography), aeg (anion exchange chromatography with gradient), gf (gel filtration) and PAGE ruler prestained protein ladder (m, MBI Fermentas) containing proteins of indicated molecular mass The arrow points at the band, which was assigned to pgm .

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Purification of the main Pgm isoenzyme of C. glutamicum , SDS/PAGE analysis of each step of the purification procedure: ce (crude extract), aec (anion exchange chromatography step wise elution), hic (hydrophobic interaction chromatography), aeg (anion exchange chromatography with gradient), gf (gel filtration) and PAGE ruler prestained protein ladder (m, MBI Fermentas) containing proteins of indicated molecular mass The arrow points at the band, which was assigned to pgm .

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques: Purification, SDS Page, Chromatography, Hydrophobic Interaction Chromatography, Filtration, Polyacrylamide Gel Electrophoresis

    Growth (circles), intracellular glc-1-P levels (bars) and glycogen content (triangles) of C. glutamicum WT (filled symbols, black bars) and C. glutamicum IMpgm (open symbols, grey bars) in the course of cultivation in minimal medium with 1% (w/v) glucose [(A) growth and glc-1-P levels, (B) glycogen content] or 1% (w/v) maltose [(C) growth and glc-1-P levels, (D) glycogen content] as sole carbon sources Glycogen was determined enzymatically as glucose liberated after amyloglucosidase treatment (two determinations per sample). Three independent cultivations were performed; the S.D. s of the glycogen contents were

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Growth (circles), intracellular glc-1-P levels (bars) and glycogen content (triangles) of C. glutamicum WT (filled symbols, black bars) and C. glutamicum IMpgm (open symbols, grey bars) in the course of cultivation in minimal medium with 1% (w/v) glucose [(A) growth and glc-1-P levels, (B) glycogen content] or 1% (w/v) maltose [(C) growth and glc-1-P levels, (D) glycogen content] as sole carbon sources Glycogen was determined enzymatically as glucose liberated after amyloglucosidase treatment (two determinations per sample). Three independent cultivations were performed; the S.D. s of the glycogen contents were

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques: Gas Chromatography

    Growth (symbols) and glycogen content (bars) of C. glutamicum IMpgm (pXMJ19) (filled triangles, black bars) and C. glutamicum IMpgm (pXMJ19-pgm) (open circles, grey bars) in the course of cultivation in minimal medium with 1% (w/v) glucose (A) or 1% (w/v) maltose (B) as sole carbon sources Glycogen was determined enzymatically as glucose liberated after amyloglucosidase treatment (two determinations per sample). Three independent cultivations were performed, data from one representative experiment are shown; the S.D. of the glycogen content was

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Growth (symbols) and glycogen content (bars) of C. glutamicum IMpgm (pXMJ19) (filled triangles, black bars) and C. glutamicum IMpgm (pXMJ19-pgm) (open circles, grey bars) in the course of cultivation in minimal medium with 1% (w/v) glucose (A) or 1% (w/v) maltose (B) as sole carbon sources Glycogen was determined enzymatically as glucose liberated after amyloglucosidase treatment (two determinations per sample). Three independent cultivations were performed, data from one representative experiment are shown; the S.D. of the glycogen content was

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques:

    Specific Pgm activity (A) and glycogen content; (B) C. glutamicum WT cells sampled in the mid-exponential growth phase from cultivations in minimal medium with various carbon sources or TY complex medium Data represent mean values of three independent determinations from at least three independent cultivations.

    Journal: Bioscience Reports

    Article Title: Inactivation of the phosphoglucomutase gene pgm in Corynebacterium glutamicum affects cell shape and glycogen metabolism

    doi: 10.1042/BSR20130076

    Figure Lengend Snippet: Specific Pgm activity (A) and glycogen content; (B) C. glutamicum WT cells sampled in the mid-exponential growth phase from cultivations in minimal medium with various carbon sources or TY complex medium Data represent mean values of three independent determinations from at least three independent cultivations.

    Article Snippet: Bacterial strains, media and growth conditions The bacteria used in this study were E. coli DH5α [ ] and C. glutamicum WT (wild-type) (strain ATCC13032; American Type Culture Collection).

    Techniques: Activity Assay

    Gas Chromatography/Mass Spectrometry (GC/MS) analysis of partially per- O -methylated, per- O -acetylated alditol acetate derivatives of purified arabinogalactan from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD . A , gas chromatograms demonstrating the presence of glycosyl linkages in purified AG. B , mass spectra fragmentation profile of peaks resolving at 11.3 min corresponding to 2-Ara f ( * ) and 3-Ara f ( ** ) and cleavage ions representing fragmentation of 2-Ara f ( * ) and 3-Ara f ( * ) located in terminal arabinan glycosyl motifs.

    Journal: Cell Surface (Amsterdam)

    Article Title: AftD functions as an α1 → 5 arabinofuranosyltransferase involved in the biosynthesis of the mycobacterial cell wall core

    doi: 10.1016/j.tcsw.2017.10.001

    Figure Lengend Snippet: Gas Chromatography/Mass Spectrometry (GC/MS) analysis of partially per- O -methylated, per- O -acetylated alditol acetate derivatives of purified arabinogalactan from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD . A , gas chromatograms demonstrating the presence of glycosyl linkages in purified AG. B , mass spectra fragmentation profile of peaks resolving at 11.3 min corresponding to 2-Ara f ( * ) and 3-Ara f ( ** ) and cleavage ions representing fragmentation of 2-Ara f ( * ) and 3-Ara f ( * ) located in terminal arabinan glycosyl motifs.

    Article Snippet: C. glutamicum strains were grown on complex medium (Brain Heart Infusion, Difco), or on salt medium CGXII , with cultivation at 30 °C.

    Techniques: Gas Chromatography, Mass Spectrometry, Gas Chromatography-Mass Spectrometry, Methylation, Purification, Acetylene Reduction Assay

    GC/MS and ES-MS characterisation of in vitro synthesized reaction products from the arabinofuranosyltransferase assays utilizing both acceptors A and B. A , GC/MS analysis and B , ES-MS analysis of products derived from assays using acceptor A and membranes prepared from C. glutamicum Δ aftB Δ aftD over-expressing Mt- aftD supplemented with EMB. C , GC/MS analysis and D , ES-MS analysis of products derived from assays using acceptor B and membranes prepared from C. glutamicum Δ aftB Δ aftD over-expressing Mt- aftD supplemented with EMB.

    Journal: Cell Surface (Amsterdam)

    Article Title: AftD functions as an α1 → 5 arabinofuranosyltransferase involved in the biosynthesis of the mycobacterial cell wall core

    doi: 10.1016/j.tcsw.2017.10.001

    Figure Lengend Snippet: GC/MS and ES-MS characterisation of in vitro synthesized reaction products from the arabinofuranosyltransferase assays utilizing both acceptors A and B. A , GC/MS analysis and B , ES-MS analysis of products derived from assays using acceptor A and membranes prepared from C. glutamicum Δ aftB Δ aftD over-expressing Mt- aftD supplemented with EMB. C , GC/MS analysis and D , ES-MS analysis of products derived from assays using acceptor B and membranes prepared from C. glutamicum Δ aftB Δ aftD over-expressing Mt- aftD supplemented with EMB.

    Article Snippet: C. glutamicum strains were grown on complex medium (Brain Heart Infusion, Difco), or on salt medium CGXII , with cultivation at 30 °C.

    Techniques: Gas Chromatography-Mass Spectrometry, Mass Spectrometry, In Vitro, Synthesized, Derivative Assay, Expressing

    Arabinofuranosyltransferase activity utilizing neoglycolipid acceptor A and membranes prepared from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD. A , Arabinofuranosyltransferase activity was determined using the synthetic neoglycolipid acceptor α-D-Ara f (1 → 3)-α-D-Ara f - O -(CH 2 ) 7 CH 3 ). The products of the assay were resuspended prior to scintillation counting (10%) and the remaining subjected to TLC using silica gel plates (5735 silica gel 60F 254 , Merck) in isopropanol:acetic acid:water (8:1:1, v/v/v) with the reaction products visualized by autoradiography. The TLC autoradiogram is representative of several independent experiments. B , biosynthetic reaction scheme of products A 1 , A 2 and A 3 formed in arabinofuranosyltransferase assays using the neoglycolipid acceptor A. C , GC/MS analysis of the partially per- O -methylated, per- O -acetylated alditol acetate derivative of reaction products obtained from assays containing membranes prepared from either C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD.

    Journal: Cell Surface (Amsterdam)

    Article Title: AftD functions as an α1 → 5 arabinofuranosyltransferase involved in the biosynthesis of the mycobacterial cell wall core

    doi: 10.1016/j.tcsw.2017.10.001

    Figure Lengend Snippet: Arabinofuranosyltransferase activity utilizing neoglycolipid acceptor A and membranes prepared from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD. A , Arabinofuranosyltransferase activity was determined using the synthetic neoglycolipid acceptor α-D-Ara f (1 → 3)-α-D-Ara f - O -(CH 2 ) 7 CH 3 ). The products of the assay were resuspended prior to scintillation counting (10%) and the remaining subjected to TLC using silica gel plates (5735 silica gel 60F 254 , Merck) in isopropanol:acetic acid:water (8:1:1, v/v/v) with the reaction products visualized by autoradiography. The TLC autoradiogram is representative of several independent experiments. B , biosynthetic reaction scheme of products A 1 , A 2 and A 3 formed in arabinofuranosyltransferase assays using the neoglycolipid acceptor A. C , GC/MS analysis of the partially per- O -methylated, per- O -acetylated alditol acetate derivative of reaction products obtained from assays containing membranes prepared from either C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD.

    Article Snippet: C. glutamicum strains were grown on complex medium (Brain Heart Infusion, Difco), or on salt medium CGXII , with cultivation at 30 °C.

    Techniques: Activity Assay, Acetylene Reduction Assay, Thin Layer Chromatography, Autoradiography, Gas Chromatography-Mass Spectrometry, Methylation

    Arabinofuranosyltransferase activity utilizing neoglycolipid acceptor B and membranes prepared from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD . A , Arabinofuranosyltransferase activity was determined using the synthetic neoglycolipid acceptor [α-D-Ara f (1 → ]3,5-α-D-Ara f - O -(CH 2 ) 7 CH 3 ). The products of the assay were resuspended prior to scintillation counting (10%) and the remaining subjected to TLC using silica gel plates (5735 silica gel 60F 254 , Merck) in isopropanol:acetic acid:water (8:1:1, v/v/v) with the reaction products visualized by autoradiography. The TLC autoradiogram is representative of several independent experiments. B , Biosynthetic reaction scheme of products B 1 , B 2 and B 3 formed in arabinofuranosyltransferase assays using the neoglycolipid acceptor B. C , GC/MS analysis of the partially per- O -methylated, per- O -acetylated alditol acetate derivative of reaction products obtained from assays containing membranes prepared from either C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD.

    Journal: Cell Surface (Amsterdam)

    Article Title: AftD functions as an α1 → 5 arabinofuranosyltransferase involved in the biosynthesis of the mycobacterial cell wall core

    doi: 10.1016/j.tcsw.2017.10.001

    Figure Lengend Snippet: Arabinofuranosyltransferase activity utilizing neoglycolipid acceptor B and membranes prepared from C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD . A , Arabinofuranosyltransferase activity was determined using the synthetic neoglycolipid acceptor [α-D-Ara f (1 → ]3,5-α-D-Ara f - O -(CH 2 ) 7 CH 3 ). The products of the assay were resuspended prior to scintillation counting (10%) and the remaining subjected to TLC using silica gel plates (5735 silica gel 60F 254 , Merck) in isopropanol:acetic acid:water (8:1:1, v/v/v) with the reaction products visualized by autoradiography. The TLC autoradiogram is representative of several independent experiments. B , Biosynthetic reaction scheme of products B 1 , B 2 and B 3 formed in arabinofuranosyltransferase assays using the neoglycolipid acceptor B. C , GC/MS analysis of the partially per- O -methylated, per- O -acetylated alditol acetate derivative of reaction products obtained from assays containing membranes prepared from either C. glutamicum , C. glutamicum Δ aftB , C. glutamicum Δ aftD and C. glutamicum Δ aftB Δ aftD.

    Article Snippet: C. glutamicum strains were grown on complex medium (Brain Heart Infusion, Difco), or on salt medium CGXII , with cultivation at 30 °C.

    Techniques: Activity Assay, Acetylene Reduction Assay, Thin Layer Chromatography, Autoradiography, Gas Chromatography-Mass Spectrometry, Methylation

    Transformation assays of FA and VAN. C. glutamicum strain F was cultured in BHI medium at 30 °C for 24 h, and 0.1 mL of each culture was transferred to 5 mL of AR medium containing 2 mM each of FA or VAN with 25 μg/mL of Km in 5 mL tubes. The biotransformation of a FA and b VAN was performed at 30 °C with agitation at 180 rpm for 72 h. The concentrations of extracellular FA ( squares ), VA ( triangles ), VAN ( diamonds ) and PCA ( circles ) were monitored every 24 h. Data represent the mean and standard error from three independent experiments

    Journal: AMB Express

    Article Title: Biotransformation of ferulic acid to protocatechuic acid by Corynebacterium glutamicum ATCC 21420 engineered to express vanillate O-demethylase

    doi: 10.1186/s13568-017-0427-9

    Figure Lengend Snippet: Transformation assays of FA and VAN. C. glutamicum strain F was cultured in BHI medium at 30 °C for 24 h, and 0.1 mL of each culture was transferred to 5 mL of AR medium containing 2 mM each of FA or VAN with 25 μg/mL of Km in 5 mL tubes. The biotransformation of a FA and b VAN was performed at 30 °C with agitation at 180 rpm for 72 h. The concentrations of extracellular FA ( squares ), VA ( triangles ), VAN ( diamonds ) and PCA ( circles ) were monitored every 24 h. Data represent the mean and standard error from three independent experiments

    Article Snippet: Genomic DNA was prepared from C. glutamicum strain F (ATCC 21420) grown in 5 mL of BHI medium using a Wizard Gnomic DNA Purification Kit (Promega, Madison, WI, USA).

    Techniques: Transformation Assay, Cell Culture

    Biotransformation of VA to PCA by C. glutamicum expressing vanillate O -demethylase (Van). a C. glutamicum strain F ( open symbols ) as a control and b strain FVan ( closed symbols ) were cultured in BHI medium at 30 °C for 24 h, and 0.5 mL of each culture was transferred to 25 mL of AR medium containing 2.5 mM VA. The bioconversion was performed at 30 °C for 48 h with agitation at 180 rpm. Extracellular VA ( triangles ) and PCA ( circles ) were analyzed every 24 h. OD 600 ( diamonds ) was monitored simultaneously. Data represent the mean and standard error of three independent experiments

    Journal: AMB Express

    Article Title: Biotransformation of ferulic acid to protocatechuic acid by Corynebacterium glutamicum ATCC 21420 engineered to express vanillate O-demethylase

    doi: 10.1186/s13568-017-0427-9

    Figure Lengend Snippet: Biotransformation of VA to PCA by C. glutamicum expressing vanillate O -demethylase (Van). a C. glutamicum strain F ( open symbols ) as a control and b strain FVan ( closed symbols ) were cultured in BHI medium at 30 °C for 24 h, and 0.5 mL of each culture was transferred to 25 mL of AR medium containing 2.5 mM VA. The bioconversion was performed at 30 °C for 48 h with agitation at 180 rpm. Extracellular VA ( triangles ) and PCA ( circles ) were analyzed every 24 h. OD 600 ( diamonds ) was monitored simultaneously. Data represent the mean and standard error of three independent experiments

    Article Snippet: Genomic DNA was prepared from C. glutamicum strain F (ATCC 21420) grown in 5 mL of BHI medium using a Wizard Gnomic DNA Purification Kit (Promega, Madison, WI, USA).

    Techniques: Expressing, Cell Culture

    Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Journal: Applied and Environmental Microbiology

    Article Title: Genealogy Profiling through Strain Improvement by Using Metabolic Network Analysis: Metabolic Flux Genealogy of Several Generations of Lysine-Producing Corynebacteria

    doi: 10.1128/AEM.68.12.5843-5859.2002

    Figure Lengend Snippet: Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Article Snippet: The strains C. glutamicum ATCC 21526 and ATCC 21543 have a requirement for homoserine and leucine and are resistant to the lysine analogue S -β-aminoethyl-cysteine.

    Techniques:

    Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Journal: Applied and Environmental Microbiology

    Article Title: Genealogy Profiling through Strain Improvement by Using Metabolic Network Analysis: Metabolic Flux Genealogy of Several Generations of Lysine-Producing Corynebacteria

    doi: 10.1128/AEM.68.12.5843-5859.2002

    Figure Lengend Snippet: Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Article Snippet: Due to their lower lysine yields and similar anabolic demands compared to C. glutamicum ATCC 21543, the strains C. glutamicum ATCC 13287, ATCC 21253, and ATCC 21526 showed slightly lower fluxes of 18.8, 21.4, and 19.9%, respectively.

    Techniques:

    Profile of batch cultivation of lysine-producing C. glutamicum ATCC 13287. (A) Volumetric carbon dioxide production rate ( Q CO2 ), concentrations of glucose and citrate, and CDM. (B) Concentrations of methionine, threonine, and lysine. (C) Concentrations of glycine, alanine, and valine. (D) Concentrations of glycerol, dihydroxyacetone, and acetate. The beginning of the lysine production phase is marked by a vertical line.

    Journal: Journal of Bacteriology

    Article Title: In-Depth Profiling of Lysine-Producing Corynebacterium glutamicum by Combined Analysis of the Transcriptome, Metabolome, and Fluxome

    doi: 10.1128/JB.186.6.1769-1784.2004

    Figure Lengend Snippet: Profile of batch cultivation of lysine-producing C. glutamicum ATCC 13287. (A) Volumetric carbon dioxide production rate ( Q CO2 ), concentrations of glucose and citrate, and CDM. (B) Concentrations of methionine, threonine, and lysine. (C) Concentrations of glycine, alanine, and valine. (D) Concentrations of glycerol, dihydroxyacetone, and acetate. The beginning of the lysine production phase is marked by a vertical line.

    Article Snippet: The same phenomenon of prolonged growth after the consumption of threonine and methionine was previously observed for lysine-producing C. glutamicum ATCC 21253, a strain directly derived from C. glutamicum ATCC 13287 and thus very similar to it ( ).

    Techniques:

    Fed-batch fermentation of violacein with C. glutamicum ATCC 21850 (pEC-C-vio1) in 3 L NBS BioFlo 115 fermenter

    Journal: Microbial Cell Factories

    Article Title: Engineering Corynebacterium glutamicum for violacein hyper production

    doi: 10.1186/s12934-016-0545-0

    Figure Lengend Snippet: Fed-batch fermentation of violacein with C. glutamicum ATCC 21850 (pEC-C-vio1) in 3 L NBS BioFlo 115 fermenter

    Article Snippet: C. glutamicum strain ATCC 21850 is an established industrial l -tryptophan hyperproducer, which made it an attractive platform since l -tryptophan is direct precursor for the production of crude violacein.

    Techniques:

    galM and araR promoter activities. (A) araR promoter activity. The 400-bp araR upstream region was fused to the lacZ reporter gene in the canonical (P araR -F) or the reverse (P araR -R) orientation. The C. glutamicum ATCC 31831 strains carrying the respective

    Journal: Journal of Bacteriology

    Article Title: AraR, an l-Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

    doi: 10.1128/JB.00314-15

    Figure Lengend Snippet: galM and araR promoter activities. (A) araR promoter activity. The 400-bp araR upstream region was fused to the lacZ reporter gene in the canonical (P araR -F) or the reverse (P araR -R) orientation. The C. glutamicum ATCC 31831 strains carrying the respective

    Article Snippet: The araA single deletion mutant strain derived from C. glutamicum ATCC 31831 wild-type strain (see Table S1 in the supplemental material; Δ araA ) was described previously ( ).

    Techniques: Activity Assay

    Effects of BS B mutation on the divergent araBDA and galM-araR operons. The relative mRNA levels of araB , araD , and araA (A) and galM and araR (B) were determined. The C. glutamicum ATCC 31831 WT strain and the AraR-binding site mutant B-m strains were

    Journal: Journal of Bacteriology

    Article Title: AraR, an l-Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

    doi: 10.1128/JB.00314-15

    Figure Lengend Snippet: Effects of BS B mutation on the divergent araBDA and galM-araR operons. The relative mRNA levels of araB , araD , and araA (A) and galM and araR (B) were determined. The C. glutamicum ATCC 31831 WT strain and the AraR-binding site mutant B-m strains were

    Article Snippet: The araA single deletion mutant strain derived from C. glutamicum ATCC 31831 wild-type strain (see Table S1 in the supplemental material; Δ araA ) was described previously ( ).

    Techniques: Mutagenesis, Binding Assay

    Expression of araE in AraR-binding site mutants. Single mutations in BS E1 (E1-m) and BS E2 (E2-m) and a double mutation in these sites (E1/2-m) were introduced into the araE promoter region in the C. glutamicum ATCC 31831 chromosome. The three mutants

    Journal: Journal of Bacteriology

    Article Title: AraR, an l-Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

    doi: 10.1128/JB.00314-15

    Figure Lengend Snippet: Expression of araE in AraR-binding site mutants. Single mutations in BS E1 (E1-m) and BS E2 (E2-m) and a double mutation in these sites (E1/2-m) were introduced into the araE promoter region in the C. glutamicum ATCC 31831 chromosome. The three mutants

    Article Snippet: The araA single deletion mutant strain derived from C. glutamicum ATCC 31831 wild-type strain (see Table S1 in the supplemental material; Δ araA ) was described previously ( ).

    Techniques: Expressing, Binding Assay, Mutagenesis

    Arabinose gene cluster in C. glutamicum ATCC 31831. (A) Location and direction of l -arabinose utilization genes ( araA , araB , araD , araE , araR , and galM ). The amplified fragments in are indicated by numbers and lines. (B and C) Nucleotide sequences

    Journal: Journal of Bacteriology

    Article Title: AraR, an l-Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

    doi: 10.1128/JB.00314-15

    Figure Lengend Snippet: Arabinose gene cluster in C. glutamicum ATCC 31831. (A) Location and direction of l -arabinose utilization genes ( araA , araB , araD , araE , araR , and galM ). The amplified fragments in are indicated by numbers and lines. (B and C) Nucleotide sequences

    Article Snippet: The araA single deletion mutant strain derived from C. glutamicum ATCC 31831 wild-type strain (see Table S1 in the supplemental material; Δ araA ) was described previously ( ).

    Techniques: Amplification

    Effects of disruption of araA on the l -arabinose-inducible gene expression. The C. glutamicum ATCC 31831 wild type (WT) and the mutants deficient in the AraR-binding site upstream of araB in the background of the wild-type strain (WT/B-m) or the araA

    Journal: Journal of Bacteriology

    Article Title: AraR, an l-Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

    doi: 10.1128/JB.00314-15

    Figure Lengend Snippet: Effects of disruption of araA on the l -arabinose-inducible gene expression. The C. glutamicum ATCC 31831 wild type (WT) and the mutants deficient in the AraR-binding site upstream of araB in the background of the wild-type strain (WT/B-m) or the araA

    Article Snippet: The araA single deletion mutant strain derived from C. glutamicum ATCC 31831 wild-type strain (see Table S1 in the supplemental material; Δ araA ) was described previously ( ).

    Techniques: Expressing, Binding Assay

    Time course metabolomics of aerobically growing wild-type C. glutamicum ATCC 31831 (black diamonds) and recombinant strains overexpressing pyruvate kinase in the absence or presence of araR deletion (31831/pCH pyk , blue triangles; Δ araR /pCH pyk , red circles). Relative abundances of metabolic intermediates in cells during aerobic growth in BT medium containing a mixture of d -glucose and l -arabinose are shown. The X-axis indicates cultivation time (h). For the profiling experiment, three strains were grown aerobically to late log phase in A medium containing 20 g/L of l -arabinose and 20 g/L of d -glucose, and the culture was then inoculated to an initial OD 600 of 0.2 into BT medium containing a mixture of d -glucose and l -arabinose as the carbon sources (15 g/L each). The cells were harvested at 11, 17, and 23 h of cultivation and subsequently subjected to metabolome analysis. Data are presented as average ± standard deviation calculated from the results of triplicate individual experiments. The concentration of erythrose-4-phosphate was below the detection limit

    Journal: Microbial Cell Factories

    Article Title: Metabolome analysis-based design and engineering of a metabolic pathway in Corynebacterium glutamicum to match rates of simultaneous utilization of d-glucose and l-arabinose

    doi: 10.1186/s12934-018-0927-6

    Figure Lengend Snippet: Time course metabolomics of aerobically growing wild-type C. glutamicum ATCC 31831 (black diamonds) and recombinant strains overexpressing pyruvate kinase in the absence or presence of araR deletion (31831/pCH pyk , blue triangles; Δ araR /pCH pyk , red circles). Relative abundances of metabolic intermediates in cells during aerobic growth in BT medium containing a mixture of d -glucose and l -arabinose are shown. The X-axis indicates cultivation time (h). For the profiling experiment, three strains were grown aerobically to late log phase in A medium containing 20 g/L of l -arabinose and 20 g/L of d -glucose, and the culture was then inoculated to an initial OD 600 of 0.2 into BT medium containing a mixture of d -glucose and l -arabinose as the carbon sources (15 g/L each). The cells were harvested at 11, 17, and 23 h of cultivation and subsequently subjected to metabolome analysis. Data are presented as average ± standard deviation calculated from the results of triplicate individual experiments. The concentration of erythrose-4-phosphate was below the detection limit

    Article Snippet: Specific growth rates and specific sugar consumption rates by recombinant strains and the wild-type strain of C. glutamicum ATCC 31831.

    Techniques: Recombinant, Standard Deviation, Concentration Assay

    Metabolic pathway of d -glucose and l -arabinose ( a ) and the chromosomal gene cluster regulating l -arabinose utilization ( b ) in Corynebacterium glutamicum ATCC 31831

    Journal: Microbial Cell Factories

    Article Title: Metabolome analysis-based design and engineering of a metabolic pathway in Corynebacterium glutamicum to match rates of simultaneous utilization of d-glucose and l-arabinose

    doi: 10.1186/s12934-018-0927-6

    Figure Lengend Snippet: Metabolic pathway of d -glucose and l -arabinose ( a ) and the chromosomal gene cluster regulating l -arabinose utilization ( b ) in Corynebacterium glutamicum ATCC 31831

    Article Snippet: Specific growth rates and specific sugar consumption rates by recombinant strains and the wild-type strain of C. glutamicum ATCC 31831.

    Techniques:

    Metabolic profile of aerobically growing wild-type strain C. glutamicum ATCC 31831. Relative abundance of metabolic intermediates in C. glutamicum cells during aerobic growth in BT medium containing either d -glucose (open columns) or l -arabinose (gray columns) as the sole carbon source are shown. For the profiling experiment, the wild-type strain was grown aerobically to late log phase in A medium containing d -glucose (20 g/L) and l -arabinose (20 g/L), and the culture was then inoculated to an initial OD 600 of 0.2 into BT medium containing either d -glucose (20 g/L) or l -arabinose (20 g/L) as the sole carbon source. The cells were harvested at mid log phase (OD 600 = 1.5) and subjected to metabolic profiling. Data are presented as average ± standard deviation calculated from the results of triplicate individual experiments. Single and double asterisks indicate p values of

    Journal: Microbial Cell Factories

    Article Title: Metabolome analysis-based design and engineering of a metabolic pathway in Corynebacterium glutamicum to match rates of simultaneous utilization of d-glucose and l-arabinose

    doi: 10.1186/s12934-018-0927-6

    Figure Lengend Snippet: Metabolic profile of aerobically growing wild-type strain C. glutamicum ATCC 31831. Relative abundance of metabolic intermediates in C. glutamicum cells during aerobic growth in BT medium containing either d -glucose (open columns) or l -arabinose (gray columns) as the sole carbon source are shown. For the profiling experiment, the wild-type strain was grown aerobically to late log phase in A medium containing d -glucose (20 g/L) and l -arabinose (20 g/L), and the culture was then inoculated to an initial OD 600 of 0.2 into BT medium containing either d -glucose (20 g/L) or l -arabinose (20 g/L) as the sole carbon source. The cells were harvested at mid log phase (OD 600 = 1.5) and subjected to metabolic profiling. Data are presented as average ± standard deviation calculated from the results of triplicate individual experiments. Single and double asterisks indicate p values of

    Article Snippet: Specific growth rates and specific sugar consumption rates by recombinant strains and the wild-type strain of C. glutamicum ATCC 31831.

    Techniques: Standard Deviation

    Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. glutamicum KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.

    Journal: Applied and Environmental Microbiology

    Article Title: Hyperproduction of Tryptophan by Corynebacterium glutamicum with the Modified Pentose Phosphate Pathway

    doi:

    Figure Lengend Snippet: Construction of low-copy-number plasmid pIK9960 containing the transketolase gene as well as the DS gene, the PGD gene, and the tryptophan-biosynthetic gene cluster. Symbols: stippled bars, C. glutamicum KY10694 chromosomal DNA fragment containing the DS gene; solid bars, C. glutamicum KY10894 chromosomal DNA fragment containing the tryptophan-biosynthetic gene cluster ( trp genes); hatched bars, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the PGD gene; cross-hatched bar, C. glutamicum ATCC 31833 chromosomal DNA fragment containing the transketolase (TK) gene; open bars, vector pCG116 (pCG1 origin), pCSEK20 (pCG2 origin), or pCS43 (pCG4 origin). Sp r , spectinomycin resistance; Km r , kanamycin resistance; Ori, origin.

    Article Snippet: Plasmids pLTK65 and pCTK60 ( ) contain the transketolase gene of the wild-type strain C. glutamicum ATCC 31833 in vectors pCLEK4 and pCSEK20 , respectively.

    Techniques: Low Copy Number, Plasmid Preparation

    Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Journal: Applied and Environmental Microbiology

    Article Title: Genealogy Profiling through Strain Improvement by Using Metabolic Network Analysis: Metabolic Flux Genealogy of Several Generations of Lysine-Producing Corynebacteria

    doi: 10.1128/AEM.68.12.5843-5859.2002

    Figure Lengend Snippet: Impact of strain optimization of lysine-producing strains of C. glutamicum on NADPH metabolism. NADPH demand (A) and supply (B) are expressed as molar fluxes related to the glucose uptake flux in five successive generations of a genealogy of lysine producers estimated via metabolic network analysis. The strain numbers represent the different generations C. glutamicum ATCC 13032 (1), ATCC 13287 (2), ATCC 21253 (3), ATCC 21526 (4), and ATCC 21543 (5). ICD, isocitrate dehydrogenase.

    Article Snippet: In comparison, the data for the other strains range between those for the wild-type strain and C. glutamicum ATCC 21543.

    Techniques:

    Statistical analysis for estimates of fluxes into the PPP and through isocitrate (ICI) dehydrogenase for C. glutamicum ATCC 13032 (strain 1), ATCC 21253 (strain 2), and ATCC 21543 (strain 3). The data shown for each strain are the results of 250 flux-independent flux estimations by a Monte-Carlo approach.

    Journal: Applied and Environmental Microbiology

    Article Title: Genealogy Profiling through Strain Improvement by Using Metabolic Network Analysis: Metabolic Flux Genealogy of Several Generations of Lysine-Producing Corynebacteria

    doi: 10.1128/AEM.68.12.5843-5859.2002

    Figure Lengend Snippet: Statistical analysis for estimates of fluxes into the PPP and through isocitrate (ICI) dehydrogenase for C. glutamicum ATCC 13032 (strain 1), ATCC 21253 (strain 2), and ATCC 21543 (strain 3). The data shown for each strain are the results of 250 flux-independent flux estimations by a Monte-Carlo approach.

    Article Snippet: In comparison, the data for the other strains range between those for the wild-type strain and C. glutamicum ATCC 21543.

    Techniques: